text:
Do 0.2 vs 0.5 ml epitubes require different PCR machines, incubators, etc?
Eppendorf themselves and many other companies list on their website two above mentioned sizes of "PCR tubes". As I understand 0.2 ml is by far more widespread and "standard" for PCR, so do they make different machines for 0.5 ml tubes or do 0.5 ml tubes fit 0.2 ml holes? Also, what about incubators and other bio equipment that holds epitubes?

Please edit the question to limit it to a specific problem with enough detail to identify an adequate answer.

I have always done PCR and qPCR reactions in 0.2ml tubes, not aware of any blocks designed for 0.5 ml, but they may exist. But there are other form factors that you have to pay attention to. Some (but not all) qPCR cyclers are made to work with low profile PCR tubes only. Some older machines can only use high profile (standard) tubes, but adapters are available. Some manufacturers have even devised multi-instrument workflows that require both. You also have to pay attention to tube color if you're doing qPCR (when in doubt, stick with clear) and skirting on 96-well plates.

The answers are both yes and no.
Many PCR machines come with the block designed to hold both 0.2 and 0.5 ml tubes (e.g. this one from Eppendorf**), or with swappable blocks (e.g. Bio-Rad C1000**), this depends on the manufacturer, the instrument you are looking at using, and what specifications you are looking for. For some machines, such as qPCR ones, there isn't any point in having 0.5 ml tube holes as almost all the assays will be done in 96-well 0.2 ml plates or 384 well plates (0.02 ml; with a different block designed to hold these plates). A large portion of whether you can use different tubes in different PCR machines is down to how the (heated) lid closes - if the block is for 0.5 ml tubes, then the lid will not go down far enough for 0.2 ml tubes, unless it is adjustable. Conversely if it is set up for 0.2 ml tubes, the lid will go down too far for 0.5 ml tubes, deforming them and potentially damaging the machine too.
Static temperature incubators, such as thermomixers (Eppendorf brand**) often only come in one tube size (e.g. Eppendorf ThermoMixer F), but there are thermomixers that come with swappable tube holders (like the ThermoMixer C), with anything between 0.5 ml - 50 ml tube holes and even plate holders.
For tube racks, it depends again - some do, some don't. Many 1.5 ml tube racks can be flipped over to expose 0.5 ml tube holes, but it depends on the brand. You can certainly buy racks for 0.2 ml tubes (usually in 96-well format), but 10 ul - 200 ul tip boxes work well for this purpose too. 0.5 ml tubes will fit into 0.2 ml racks, but will be unstable and you won't be able to pack the tubes as densely as 0.2 ml tubes.
** I have no affiliation with Eppendorf, Bio-Rad, nor any other brand and do not endorse or imply endorsement through this post. I also do not I sell or distribute any products - I'm simply an end-user.

Here's a top-down photo of the heating block from an Eppendorf 6331 Nexus Gradient MasterCycler (source):

Most modern Eppendorf thermal cyclers have a similar block.

Are those super little ones for 0.02 ml tubes @bob1 mentions?

@beardeadclown no. count the super-small holes and you will notice that they are way less than 384. Plates with smaller volume per tube are higher density since they split the positions into 4 smaller ones when going from coarse to finer

@masterX244 I'm sorry, I didn't get any of that. What's the significance of number 384? Plates I assume mean tube holders, coarse being .5ml and finer being .2ml, and density means holes per area, but I can't see what positions do those super-small holes split into 4 smaller ones. Also, what are those 6 flat holes for?

@beardeadclown 384=4x96. Each "position" gets split into 4 smaller ones when raising plate density (except when going from a Basin plate which is a 1x1). Stepping is 6, 24, 96, 384,1536 on the standard plates which all got the same size. And i guess that the 6 flat holes are due to technical reasons.

@beardeadclown I imagine the smaller holes are to accommodate expansion and contraction of the block as it is rapidly heated / cooled. I'm not sure what the six larger holes are for, but I don't think they are meant to hold tubes

@masterX244 I think I'm starting to understand. Is there any freely available resource where I can learn more about PCR standard plates, tubes, their dimensions, design and technical specifications?

You can start by browsing brochures from Eppendorf and other manufacturers.


text:
How precise does a 1:1 ratio PCR sample mixture (microliters) have to be for the liquid amount ratio to not have any effect on results
Amount of liquid:
5 micro liters master mix
5 micro liters of patient sample

Please clarify your specific problem or provide additional details to highlight exactly what you need. As it's currently written, it's hard to tell exactly what you're asking.

Welcome to the site - can you please let us know what sort of PCR you are doing. Regular PCR with a gel as end-point, it might not matter too much (i.e. you just want a band), but for qPCR it will make all the difference.

If your pipets are calibrated and your technique is good, you shouldn't have issues with those volumes. Using a p10 instead of a p20 might reduce pipetting error a bit. Low-binding tips can help too, but marginally. If running reactions in duplicate or triplicate you can mix them all in one tube first, and dispense the 10 ul reactions out from that; just make them around 10% or so bigger than what you need to account for dead volume, and wet your tips before dispensing.

Pipetting PCR reactions should be done as precise as possible - since relatively small deviations can effect the reaction. Pipetting 4 instead of 5µl results in a 10% error which for example leads to a higher magnesium concentration which may effect the reaction efficiency.
I would try to use bigger volumes (25µl is pretty common for PCR reactions) to minimize such effects, but of course smaller reactions save material and costs. You could run a few tests to see how big the effect is for different reaction volumes and if the possible error is acceptable for you.

Could you clarify more on what would be an acceptable range of deviation in that it wouldn't effect the reaction or provide me please with any reference I could view? For example if I had a total mixture of 20µl.

@Nicole you can work out the error by yourself fairly easily - it's simply your pipetting volume error divided by the expected total volume (e.g if you had a 1 ul error in expected volume 20 ul - 1/20 = 0.05 = 5%)

thank you all for the input

source: Stack Exchange

license: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/4.0/

sort: votes


text:
What benefit do cardiomyocytes accrue by requiring calcium induced calcium release (relative to skeletal myocytes)?
According to 2 sources I've read, in contrast to skeletal myocytes, cardiac myocytes need calcium to diffuse in to result in contraction. One source says that they need large amounts of calcium to diffuse in through T-tubules as the cardiac sarcoplasmic reticulum is not as well developed as the skeletal sarcoplasmic reticulum, whereas the other source says that a small amount of calcium diffusing inwards is required to activate ryanodine receptor channels in the cardiac sarcoplasmic reticulum which then release large quantities of calcium which result in contraction. Regardless, what benefit do cardiac myocytes gain over skeletal muscles by being regulated by extracellular calcium? My best guess is that it helps prevent the possibility of tetany, but I am unsure.

It would be helpful to include specific links to the sources you refer to. You might also have more luck with the Medical Sciences Stack Exchange. https://medicalsciences.stackexchange.com/

all_licenses: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/4.0/


text:
How much is known about dinosaur DNA condensation and crosslinking?
The conclusion section of Bailleul et al. (2020) Evidence of proteins, chromosomes and chemical markers of DNA in exceptionally preserved dinosaur cartilage is:

The identification of chemical markers of DNA in Hypacrosaurus suggest it may preserve much longer than originally proposed [30,31]. Even though it is clear that contamination does exist in fossil material and complicates identifications of original organic molecules, it can be accounted for with proper controls. Contamination is not a plausible explanation in this case, and to this date, the possible preservation of original proteins and DNA in deep time has not been convincingly eliminated with data. Although extensive research and sequencing is required to further understand DNA preservation in Mesozoic material, along with its chemical and molecular alterations, our data suggest the preserved nuclear material in Hypacrosaurus was in a condensed state at the time of the death of the organism, which may have contributed to its stability. We propose that DNA condensation may be a favorable process to its fossilization. Additionally, as was suggested for protein fossilization [20,45,46], crosslinking may be another mechanism involved in the preservation of DNA in deep time.

The last three sentences highlight that "the preservation of DNA in deep time" may be aided by both DNA condensation and crosslinking.
Wikipedia's DNA condensation; in eukaryotes links to Chromatin
If I had to bet I'd say dinosaurs had chromatin but I could not defend it. If one of the conclusions of this paper was that the DNA was in a condensed state, then it sounds like they are not taking it for granted either.
Is this paper the full extent of what's known about DNA condensation in dinosaurs?
Together with condensation, crosslinking is mentioned in the conclusion, but a text search for "crossl" finds no other instances. As it seems closely related to condensation but there's no further information I don't see how to break that out as a separate question yet.
Question: How much is known about dinosaur DNA condensation and crosslinking?
Potentially related and/or helpful:

Zheng et al. (2021) Nuclear preservation in the cartilage of the Jehol dinosaur Caudipteryx
Wikipedia: Hypacrosaurus; cells

Biology SE:

Was 14,000+ year old DNA "laying around in cave dirt" protected from degradation, or is it just naturally this robust?
Roughly speaking how stable are short sections of single-stranded RNA in exposed environments compared to double-stranded DNA?

I'd say about as much as is known in birds...

@bob1 actually known?

Well, no, not actually known, given that there's 65 My between then and now, but I doubt such important mechanisms have changed much in the intervening time.

@LifeInTheTrees It can not fail if it does not try. It does not try because as far as I can understand (not being a molecular biologists) they simply did not search for DNA. This is in fact what makes the paper so intriguing and tantalizing! However, my question is not about this paper specifically (it's provided as evidence of prior research and what got me interested in this. There is likely more known on the topic (I have linked to a second paper just as an example) but probably not much more, and so I've simply asked "How much is known...?"

Of course dinosaurs had chromatin. Histones — a key constituent of chromatin — are present in eukaryotes as primitive as yeast. I am not aware of any way to establish the condensation state of fossil chromatin, nor would I imagine it to attract much interest, but I may be wrong.

phylogenic bracketing is your friend, if crocodilians and birds both have it non-avian dinosaurs would have almost certainly have had it.

@John I think a short answer based on that would be well received. It's not exactly "known" in the same way that we know what their bones look like, but I think if you qualify it, it's certainly helpful! I'd rather have phylogenic bracketing as my friends than crocodiles and dinosaurs :-)

@uhoh Phylogenetic bracketing was what my first comment was about (more or less) - look at closely related species and work from there. I know hardly anything about DNA condensation, so I won't attempt an answer myself at this point.


text:
Lab device to periodically, automatically transfer zooplankton between containers
I need a device to periodically draw a sample of transfer small aquatic organisms from one container to another without crushing/killing them.  Each sample would be on the order of 10ml.
My sketch of an idea is below.  It would use a pump to pull the sample from the source container into a reservoir through one check valve and pump it into the destination container through another.
I believe I can build it out of a peristaltic pump and commonly available check values, , but I'm worried that zooplankton won't fit through the values.
I'm also curious if there's a better and/or off-the-shelf solution for doing this kind of thing.

As if feeding predators in the second tank? Perhaps a "Fraction Collector".

There are multiple kinds of check valves such as ball valves.

@DKNguyen I've spent some time looking and I can't find any that useful for the small volumes I need other than those used in aquariums (which might work).  I'm new at this, so maybe I don't know which suppliers to check?  Any suggestions?

@PolypipeWrangler Yes, that's exactly what I'm doing.  The fraction collectors I see online don't look suitable.

Mcmaster Carr maybe?

If zooplankton are small, a dripping tap might work. Or in the predator tank have a screen (permeable to zooplankton only) and let them live there.

Please clarify your specific problem or provide additional details to highlight exactly what you need. As it's currently written, it's hard to tell exactly what you're asking.


text:
What Produces the Postive Potenial in the lumen of the thick ascending loop of Henle?

In the Thick Ascending Loop of Henle, Paracellular diffusion of certain Solutes like magnesium and calcium takes place. Such diffusion is a result of the positive lumen potential.
Looking at the image above from "Berne & Levy physiology" both sides should produce similar charge when cancelling charges out. I don't understand what exactly produces the positive potential on the lumen side.
I believe the caption attempts to explain it referring to differences in conductivity between the apical and basal membrane but I couldn't understand how they came to the conclusion.

Don't pay attention to the stoichiometry of the pumps, that will only fool you! The charge is due to passive movement through open channels. This is shown as the "yellow tubes" with dotted lines in the diagram. Students of neuroscience are often confused by a very similar thing, as the resting potential of neurons (as well as voltage during action potentials, for that matter) is determined the same way.
The explanation is in the caption text, quoting from your figure:

Because the apical membrane is conductive primarily to K+, the apical membrane voltage is more negative than the basolateral membrane voltage, which is conductive to K+ and Cl-

The other missing piece of information is that the outside space is (relatively) low in potassium. That means that, given an available path for potassium to diffuse across the membrane, there will be a net flow of potassium out of the cell. That flow causes a negative charge to develop inside, because the potassium diffusing out is positively charged. Very few ions have to actually move to develop a biologically relevant potential (see explanations in other answers involving neurons: https://biology.stackexchange.com/a/76167/27148 and https://biology.stackexchange.com/a/57066/27148), so you can treat the ion concentrations as unchanged (and they will be, out to a fraction of a percentage point) despite this movement.
At some voltage, the negative charge inside the cell will counteract the movement of potassium, so it's not like this is a constant deluge of potassium flowing out, it's more of a maintenance trickle. You can calculate this "reversal/equilibrium voltage" using the Nernst equation.
When you have multiple ion species involved, the equation to use is the Goldman equation which is related to the Nernst equation but involves a relative weighting of different ions by their concentrations and permeabilities. As in the textbook description, adding chloride conductance makes the basolateral membrane less negative than if there was only a potassium conductance, because chloride moves out of the cell as the inside becomes more negative.

But what makes the lumen potential positive relative to the extracellular space on the basolateral side? as shown in the image, the paracellular diffusion is driven by that potential difference. yes chloride exits on basolateral side, but looking at the rest of the transporters it seems like the charges should balances out on each side of the cell. Am i missing something?

@OmarShahaltough Read the explanation in my answer; don't pay attention to the movement shown in the pumps and co-transporters, those don't matter.

im sorry if i misunderstood but im not seeming to understand two things: 1. the relevance of the potential inside the cell when it comes to the potential outside of the cell, and 2. What exactly does "makes the basolateral membrane less negative mean"?

@OmarShahaltough All potentials are relative; the potential inside the membrane and potential outside it are the same, just opposite sign. The apical membrane (left side) has an electrical gradient that is more negative inside, so pulls positive charge in. The basolateral membrane is the one on the right hand side of the diagram; since chloride can move freely on that side, there isn't the same negative gradient into the cell. If you sum these in series, the net result is a pumping of positive ions from the left side through to the right (paracellular diffusion).


text:
How can a small bug dart around in a slug's slime?
Last year I saw a large slug in the garden, and when I looked closer there was a small creature moving around rapidly, and I mean really fast (hence my use of the word "dart" in the question title - it was no exaggeration!), on the slug's surface below the slime.
What could this creature have been, and how on Earth could it have moved so fast through the slug's sticky slime?

Welcome to Biology.SE. Please take the [tour] and then go through the [help] pages starting with [Ask] questions effectively on this site and [edit] your question accordingly. In particular, your question isn't clear. To identify an organism we need: a detailed description (preferable including clear photos), location, size, and appropriate tags. Also try not to leap to conclusions (e.g. that this was a parasite).

Most likely a mite, I have seen these before.
They can move (swim) faster than the slug.
Probably Riccardoella oudemansi since it is the only slug mite I can find a mention of.
Article on similar snail mite below.
http://en.wikipedia.org/wiki/Riccardoella_limacum


text:
Identification of odd jellyfish-like creature in the Mediterranean
My parents encountered this unknown creature on a beach in Karpathos, Greece.
None of the locals have seen it before, it may be a species that has moved further north due to ocean warming.
Any ideas what it is?

@tyersome - Thank you for migrating to the proper page.

It seems to be a "Hula skirt siphonophore" - Physophora hydrostatica

Physophora hydrostatica, also known as hula skirt siphonophore, is a
species of siphonophores in the family Physophoridae.1

https://en.wikipedia.org/wiki/Physophora_hydrostatica

Nicely done. Distribution is in the Mediterranean and much of the Atlantic, so that fits. There are some nice photos of it here


text:
What kind of yellow flower is this?
Shot in a garden in Haliburton, Ontario, Canada on August 6th, 2020. Flowerhead size approx. 3" diameter across.

It would be helpful to show the flower-head from a bee's eye view, perhaps the leaves too if you can.

Can unequivocally say Asteraceae from the picture. Beyond that, without more information, very hard to say. Likely a garden (cultivated) plant rather than  a wild species.

We serious gardeners call that a DYC: a damned yellow composite, because there are so damned many of them. Head on photo required, but I'd guess it was a single heliopsis. Leaves would help, too.

It's definitely heliopsis, probably Heliopsis helianthoides.


text:
Mechanism of the calcium-activated protein, Aequorin
In Aequorin, coelenterazine acts as the luciferin, producing light in the presence of calcium and oxygen.
It is unclear to me what part of the protein structure actually catalyses this reaction after the conformational change, and I do not know quite where to begin to find this information.

My edit of your question was to make the title more general, informative and consistent with the actual question, to separate the background from the actual question, and to soften the final remarks. I do not think I have changed your meaning, but do say if I have.

The scientific question
The poster is interested in how the fluorescent protein, aequorin, catalyses the production of light. He mentions the requirement for calcium and oxygen, the role of coelenterazine as a “luciferin”, and is aware of the fact that the protein undergoes an conformational change. He specifically asks

“what part of the protein structure [my emphasis] actually catalyses
this reaction after the conformational change”

I think this is best answered — and will be more useful to those who are not familiar with this protein — by first considering the overall process, and then looking at what is known about changes in structure and how this can be interpreted.
The sequence of event in aequorin fluorescence
Fluorescence of aequorins such as aequorin or obelin occurs through a cycle of events:

The protein (apoprotein) non-covalently binds the organic prosthetic group, coelenterazine, and molecular oxygen
Coelenterazine is ‘activated’ by oxidation to 2-hydroperoxycoelenterazine†
Calcium ions bind to the apoprotein
Activated coelenterazine is converted to coelenteramide and molecular carbon dioxide with concomitant release of blue light
Coelenteramide and calcium ions are released from the protein

I would imagine that coelenteramide would be converted back to coelenterazine by some process, rather than being degraded, but I have not been able to find information on this point.
†In the papers I looked at for this answer I did not encounter information on this, albeit important, aspect of the topic. No doubt such information exists.
The substrate and effector binding sites on the protein
Our knowledge of the molecular details of catalysis by proteins depends on determining structures in which the protein is ‘frozen’ at one particular stage of a reaction. In this case it is not too difficult to crystalize the protein bound to either coelenterazine or to calcium ions, but the reaction and release of these molecules hinders finding a stable structure with both bound. The structures below from the work of Liu et al. show obelin, first with bound coelenterazine, and then with bound calcium ions, although here the coelenterazine has been converted to coelenteramide. This is useful for seeing the relative positions of ions and prosthetic group within the apoprotein.

It can be seen that the three calcium ions (red balls) are bound to similar helix–loop–helix domains called EF hands which are also found in other calcium-binding proteins. In fact, the molecule has pseudo-symmetry with four such domains, but one of them does not bind calcium. The large binding site for coelenterazine is in the centre of the molecule and involves parts of the helices of the EF hands. Because of the cartoon representation, which ignores the side-chains, the binding site appears much more open than is the case.
Conformation changes on binding calcium ions
Let us consider this first from a general standpoint. Because calcium ions bind to the amino acid side-chains of EF-hands through many polar interactions and hydrogen bonds, there is a structural difference between the bound state and the unbound state, where different interactions of these side-chains will occur. Such structural changes at the calcium-binding sites are accompanied by structural changes at distal positions in the protein, including the site at which a substrate binds. This, in turn, can affect the conformation of the substrate, facilitating its reaction.  Although it may be most appealing to consider the effect of the binding of calcium or other effectors as initiating a concerted chain of events, it is perhaps more often regarded as affecting the position of equilibrium between two alternative conformations of a protein, differing only slightly in thermodynamic Gibbs free energy. This latter relates more easily to the comparison of two crystal structures of a protein, where there are “before” and “after” situations to compare.
In the case of obelin, the two structures (above) show anticipated, although relatively modest, structural differences at the calcium binding site. I do not reproduce them here as they are best inspected with a pair of stereo glasses in a printed copy of the paper Liu et al.. The same is true for the changes in the substrate binding site, but it can be seen from the figure above that there is a change in conformation of the substrate after carbon dioxide is released.
The two-dimensional comparison (below) is pertinent as it shows (as does the work of others) that there is a particularly marked change in the interaction of a specific tyrosine and histidine residue (Tyr190 and His175 in obelin, here, and Tyr184 and His169 in aequorin and that of Tyr138 (Tyr132 in aequorin) at the other ‘side’. Changes in bonding to water molecules are also seen (discussed below).

[From Liu et al. (2006) Comparison of the hydrogen-bond network in the binding cavities of obelin before (A) and after (B) discharge of calcium ions.]
How does the conformational change cause bioluminescence?
The mechanism of bioluminescence is outside my sphere of competence, and ideas on how this is triggered in aequorin are based on a range of other experiments. However the following may be of interest:
In 2000 Head et al. wrote, regarding aequorin:

Displacement of the helices flanking the C-terminal loop would be
expected to disrupt the hydrogen bonding of the loop, resulting in
relocation of the side chain of Tyr 184 [Tyr190, above]. This is in
turn would disrupt the hydrogen bonds to His 169 [His 175, above] and
the peroxide. No longer stabilized, the peroxide would be free to
attack the carbonylic C3 to initiate the light-emitting reaction.

Liu et al., writing in 2006, elaborated on this by including a role for the water molecule (W2), the change of position of which they had observed (above).  They proposed the following reaction scheme:

How does a physical scientist go about finding — and understanding — information on proteins?
In my original answer I focussed on how to find information about proteins. That I retain in a more condensed form below. It appears that the poster has a strong background in the chemistry relating to this work, but has had little contact with proteins. As someone whose first degree was in chemistry and then moved into molecular biology, I sympathize. However, although one’s chemistry makes it easy to understand the chemical principles in protein chemistry, one still has to become familiar with the structure of proteins and accustom oneself to what may seem a strange world of weak interactions and non-standard environments. There is no avoiding the need to read at least the chapters on proteins and enzymes in a solid text like Biochemistry by Berg et al., and for this purpose any old second-hand edition will do.
With this background one can, of course, use a general search engine or Google Scholar to search for information on a particular protein. However when these methods fall short I usually go to the Protein Data Bank and search for the protein involved. In this case I used the two terms, aequorin and coelenterazine. This gave me a choice of different structures. I browsed through them, had a look at the 3D views, and then tried to find a relatively recent structure which had a publication associated with it. In this case I chose 7O3U.pdb, which has a related 2022 paper in Protein Science that seemed to discuss the mechanism and active site. From there I worked back to the two papers which are cited in this answer.

Thank you for your response. I am not particularly familiar with protein classification, as my background is more on conventional chemistry rather than biochemistry. Not exactly savvy with the Protein Data Bank. Seems like a massive data load that makes finding the catalytic moiety hard. If anyone else with a better background in this wants to give it a run, feel free.

I have now addressed the scientific question and edited my general advice on finding info about proteins. I would stress that the structure of aequorin is not particularly complex, although the reaction(s) it catalyses and the way it does so are. To go further than this answer you will need to read the original literature, and to do that you will need to have mastered at least the basics of protein chemistry.


text:
How much charge is moved for one heart beat?
Roughly, how much charge cycles (in and out of our heart cells) for each heart beat?
I am just looking for the total number of Coulombs, but, since all charge can
apparently be
accounted for with just $Na^+$, $K^+$, $Cl^-$, and $Ca^{2+}$ itemization,
it would be great if you can break it down by these ions.
My crude estimation comes from some $1\mu F/cm^2$ neuron wall capacitance
references, my guessed $10cm^2$ wall area (though the heart is bigger, I just
want the neurons), and the often-quoted $70mV$ action potential.  This gives $0.7\mu C$
(which is 4 trillion singly-charged ions).  But, I suspect that the true value
is much higher based on some other animal neuron papers that I found.

Most of the heart's electrical activity is in muscles, not neurons.

@BryanKrause I might have used the term "neuron" too loosely.  I am actually interested in all of the heart's electrical activity which factors into ECG waveforms.  So, feel free to change "neuron" to "muscle" in my question (unless the electrical activity you mention somehow gets muted in ECG data...and then it would of course be nice to explain the cause for this muting in any answer).

That's going to be quite the venture... the physics are incredibly complex, because while you generate voltages across membranes, these voltages are aligned in all sorts of directions and the vast majority cancel each other out when measured at a distance. Why do you need or want to know this? The reason you can measure an EKG at distance is because of spatial organization in the depolarizaton/repolarization of the heart: the top goes first. Bit of that discussed here: https://biology.stackexchange.com/questions/90434/in-the-pr-segment-of-an-ecg-how-come-theres-no-current-flowing/90492#90492

See also https://meta.stackexchange.com/questions/66377/what-is-the-xy-problem

@BryanKrause My question did not ask about the full EKG interpretation because I know it's complex.  I just want to know the total charge that goes through neuron/muscle membranes in the heart for each heart beat (not the directions you point out...it seems like you are pushing this into an xy problem).

I'm trying to figure out why you need this information, though; what do you plan to do with it? That's the XY part. Since you mentioned you were interested in EKG, it seems relevant that answering the question as asked won't really tell you anything about EKG measurement.


text:
What determines the maximum frequency limit that humans can hear?
I learned that the human ear doesn't hear sounds outside the range of 20-20,000 Hz.
I can understand that sounds below this range are so weak that they don't affect the ear.
But why do sounds above this range have no effect on the human ear?
The reason I'm asking is that I'm going to teach a class on hearing and sound waves, but I'm confused about the direct reason of the lack of perception of ultrasonic waves - is there a barrier in the ear or what is the direct case of blocking of hearing for these waves?

Welcome  Abd-Elaziz Sharaf. This is not simply a Q&A site, here we are expected to show the results of our own investigations into a subject here, you can [edit] to tell us what your searches found and where the sticking points are. See our section on how to ask then you can [edit] your question to include your researches.

It is perhaps easier to answer the question by rephrasing it as 'What determines the frequency range that an ear can hear'?
In order for a sensory organ to respond to sound, it must be set into motion by the sound. This is usually achieved through some support structures connected to the ear. In mammals, this includes the outer ear, the ear canal, the eardrum, the middle ear and complex structures in the inner ear (the organ of Corti). Generally speaking, when these structures are very small, relative to the wavelength of the sound, they will not be moved sufficiently by the sound and therefore not respond to the sound. When they are very large, again relative to the wavelength of the sound, they cannot follow the rapid oscillations of the sound due to their inertia and therefore do not respond to the sound. Therefore, in general, large ears on large animals tend to mean better hearing at low frequencies, whereas small ears on small animals tend to mean better hearing at higher frequencies. The upper hearing limit of humans is determined by the mechanical properties of especially the middle ear and is likely a reflection of specializations to improve hearing at lower frequencies, required for hearing and understanding speech.
Many other mammals, typically smaller than humans, have much better high-frequency hearing than humans. In almost all cases, however, this is at the expense of poorer hearing at lower frequencies, where we have good hearing.

Welcome and thanks for your answer. You say: 'large ears on large animals tend to mean better hearing at low frequencies, whereas small ears on small animals tend to mean better hearing at higher frequencies.' I reckon you mean the inner ear, and specifically the length of the cochlea? This webpage from Van der Bilt uni doesn't support that claim. Further, why would mammals have evolved ears designed for low-frequency hearing? Bats are a big group that have exquisite high-f hearing

The length of the cochlea basically limits the the frequency range, assuming the frequency distribution is about the same across species (not entirely true either, given that the Greenwood map differs across species). The upper frequency limit is determined by factors other than the inner ears length and independent on the size of the animal, afaik.

There are several scaling phenomena at play here at the same time. In general, large animals have large sound producing structures and therefore are better at making loud sounds at low frequencies. At the same time, they also tend to move over larger distances than smaller animals and therefore benefit from the longer transmission of the low frequencies over the higher frequencies. They also have larger middle ears and inner ears, so it all goes hand in hand. In general.

There are always exceptions, and bats and dolphins, have extremely well-developed high-frequency hearing. For dolphins much better than what you would expect from their size alone. The reason is echolocation, because they benefit from the higher resolution they can get by using sound with small wavelengths (high frequencies) to 'see' finer details. Dolphins tell us that it is possible for mammals much larger than humans to have ultrasonic hearing. That we do not have ultrasonic hearing tells us that in our evolutionary history, ultrasonic hearing did not increase survival of our ancestors.

It is generally agreed that 20 kHz is approximately the upper human acoustical hearing limit through air conduction. The reason for this is debated, but the transfer function of the ossicle chain in the middle ear is a suspected to determine the upper limit of audible sounds to 20 kHz (Hemila et al., 1995). Indeed, by means of bone conduction we can hear up to 50 kHz, and values up to 150 kHz have been reported in the young (Pumphrey, 1950).
Others, however, have reported that it is not only the middle ear structures, but mainly the tonotopic organization together with the finite length of the inner ear (the cochlea) that determines the upper frequency range. In mammals, tonotopy arises from a number of components, namely (1) the spatially graded BM stiffness that determines passive tuning of the basilar membrane (BM) and (2) the active frequency tuning of the BM due to auditory-nerve fibers innervating adjacent inner hair cells that sharpen up the tuning curve. In turtles, however, tonotopicity results largely from (3) the spatial gradient of the electrical properties of hair cells. Whatever the mechanism, tonotopy restricts the audible frequency bandwidth by limiting the characteristic frequencies of the auditory-nerve fibers innervating the extreme apical (low frequency) and basal (high frequency) regions of the cochlea (Rechiro & Temchin, 2002)
Your notion that

I can understand that sounds below this range are so weak that they don't affect the ear.

Is partly inaccurate, because you mix up sound level and frequency. It is, however, also partly true, because (1) large-amplitude, low-frequency stimuli may become audible at high intensities due to the high-frequency tail of the frequency tuning of inner hair cells (Smith et al.,1987) and (2) because the human ear is most sensitive to mid-range frequencies, increasing the amplitude of low-frequency sounds through cochlear curvature can affects low-frequency hearing limits.
References
- Pumphrey, Nature 1950; 166: 571
- Hemila et al., Hear Res 1995; 85: 31-44
- Smith et al., AJA (1987); 29: 125-38
- Rechiro & Temchin, PNAS; 99(20): 13206-10
Further reading
- Can humans perceive sounds above 20 kHz?

Wow... that's informative... is there any diagram descriping those 3 steps ? - for my note I was taking about physical wave energy but I'm happy with your notice.. I think @Jakob discriped physically and you did physiologically but I can't imagine your description clearly

@Abd-ElazizSharaf which 3 steps?

stimuli - ear sensitivity factors - inner hair cells affect


text:
What kind of tree has leaves like this?
Shot taken in Spring, on May 22, 2020 in a forest in Haliburton, Ontario.

Your other question also shows a Beech.

Please see the guidance at https://biology.stackexchange.com/tags/species-identification/info for species ID questions and use that tag when you ask for ID.

This is a beech tree. From the leaves only it is hard to say which species it is exactly, but given your location it is probably the North America Beech, Fagus Grandifolia.
See this leave from the second Wikipedia article for comparision:


text:
Which cancer cell lines do not increase glucose uptake in comparison with their wild type analogues?
I heard that some cell lines which already have a high glucose influx do not increase it upon altering to a cancer phenotype. However, I cannot find any supporting references.

Where did you hear?

Welcome to Biology.SE. This is not simply an answer site, but instead a site that promotes self-learning with some expert help. Consequently, questions that show little or no prior research effort are off-topic on this site as are "homework" questions unless you have shown your attempt at an answer. Please [edit] your question and tell us where you've looked for answers, what you do know about the topic, and where exactly you still have questions. Please take the [tour] and consult the [help] starting with [ask] for details.


text:
What insect is this? (Switzerland)
I took these photos in June in Zurich, Switzerland. The insect struck me as fairly large; I'd say the body was probably 2-3 cm long unless my memory is playing tricks on me. It sat just like this on a paved road. When I got close to take photos, it froze completely and I could take photos for minutes without it so much as twitching. Whenever I was farther from it, it would turn and walk a bit, but it seemed in no hurry to go anywhere.

See also this post.

Ah, yes. The differences in the antennae compared to some photos I could find threw me off; turns out that just means she's female. I think that's very likely a Ctenophora ornata.


text:
In human anatomy what is the purpose of measuring shoulder abduction and shoulder adduction separately if they cover the same range of motion?
In human anatomy what is the purpose of measuring shoulder abduction and shoulder adduction separately if they cover the same range of motion?
For example, this video
Goniometry Shoulder Abduction & Adduction  uploaded by Kelly Watson
https://www.youtube.com/watch?v=WgTu6HsZZTE
mentions measuring shoulder abduction and adduction but it seems to me like maybe One would be something like 180 minus the other.
In the case of measuring knee extension and knee flexion that situation doesn't exist as extension is 0 degrees(leg straight), and possibly a few degrees if there is hyperextension.  And flexion is around 0(leg straight), to  150 degrees. So degrees of flexion and degrees of extension don't overlap. You can't substitute one for the other.
In the case of hip abduction and adduction, it's also the case that they don't overlap. For example this really clear video
Hip abduction and adduction ROM with a goniometer By MCCCPTAP
https://www.youtube.com/watch?v=CjotehYttpE
Shows where 0 is, and how hip abduction is one way from there and hip adduction is the other way, so there's no overlap. And it's useful to have a measure of each. They aren't interchangeable.
So, the shoulder abduction and shoulder adduction one seems strange to me.. What is the purpose of measuring each, if they overlap. And if they do overlap, then why the inconsistency if comparing shoulder abduction and adduction, to hip abduction and adduction?

interestingly this video https://www.youtube.com/watch?v=ih_OZB_ciT4 mentions that very rarely would you measure adduction.. unless they have a problem with their arm being at the side. But then wouldn't that still be within the abduction range? so still doesn't quite answer it

To answer this question of "In human anatomy what is the purpose of measuring shoulder abduction and shoulder adduction separately if they cover the same range of motion?"
Before looking at some example images of video clips showing movement I think it's easier to consider wrist flexion/extension, and knee flexion/extension, as the concepts are very analogous.
In the case if wrist flexion and wrist extension,  0 is neutral. And the wrist can move maybe 70 degrees either way.
In the case of the knee,  0 degrees is full extension. Some people have a few degrees of hyperextension. And basically the movement people do in a gym is moving through degrees of knee flexion, on a machine known as a knee extension machine.
So if we apply that to the case of the wrist, when people say that wrist extension typically has a ROM/range of motion of 70 degrees, they mean when measuring from 0.  A person could do wrist extension from a position of wrist flexion, and do e.g. 140 degrees of wrist extension.
So, degrees of extension and degrees of flexion do not overlap.
But the terms extension and flexion do not assume/necessitate starting from a particular position  of extension or flexion. They could start from any position of extension or flexion or neutral.
In the context of measurement with a goniometer, and measuring a person's range of motion, then it suggests starting from 0/neutral. But otherwise, just talking of the movements of extension and flexion, it doesn't require starting from zero.
So to answer the question of "In human anatomy what is the purpose of measuring shoulder abduction and shoulder adduction separately if they cover the same range of motion?"
If each starts from zero, then they don't cover the same range of motion.
If measuring a person's ROM/range of motion, as physiotherapists do, then one would  start from zero.
If not measuring a person's range of motion, e.g. if just interested in the movement then the movements can cover the same range. One might extend from a position of extension. One might extend from a position of flexion, and from there either stay in flexion, or go into extension.  Likewise one might flex from a position of flexion or extension or neutral/0.
Like forearm supination can start from a degree of pronation and finish at a degree of supination, or start from a degree of supination and finish at a degree of supination, or start from a degree of pronation and finish in a degree of pronation. A "long" forearm supination movement not measured from 0, would start from a high degree of pronation and finish at a degree of supination.

I think the term arm abduction/adduction normally refers to vertical abduction/adduction, but i'll mention horizontal abduction/adduction too.
A screenshot from this video on youtube, man with red dot on his shoulder, shows the furthest degree of (vertical) shoulder adduction.

That's  consistent with what I heard from somebody that told me "normal (vertical) shoulder adduction motion is 30 to 50 degrees. Normal range of motion for shoulder (vertical) abduction is 150 degrees. "
There is an image from https://www.anatomyexhibits.com/  showing 0-180 degrees of vertical shoulder abduction.
Without reproducing it here , it's like

And they have an image showing 0-30 degrees of shoulder vertical adduction
which you can imagine with a protractor showing 0-30 degrees beyond 0. So  the arm starts by the side at 0 (neutral position), then moves across the body like the first pic(the man with the red dot on his shoulder).
The full movement of adduction would go from a high degree of abduction, down to 0 degrees of abduction / 0 degrees of adduction, and through to a high degree of adduction(as far as the arm goes for adduction to the end of the movement).
So likewise, with the hips , degrees of hip adduction are different to degrees of hip abduction. Though the movements of abduction or adduction can pass through degrees of one or the other or both. And that applies with wrist extension and flexion too. But for measuring a person's ROM you'd start from 0.
And I notice that the anatomyexhibits link specifies, vertical shoulder abduction/adduction.   Horizontal shoulder Abduction/adduction  would be in the transverse plane (I have noticed a difference though where some eg anatomyexhibits, measure horizontal shoulder abduction/adduction with 0 out in front, whereas others measure it with 0 to the side).
e.g. this link https://www.researchgate.net/figure/Shoulder-horizontal-adduction-A-start-position-B-end-position_fig1_309743960
has horizontal shoulder adduction (or no doubt abduction), with 0 out to the side.

whereas anatomyexhibits.com has 0 out in the front
--
This 7sec video saying it shows shoulder adduction.. youtube.com/watch?v=2N6WhuGDZRs is showing it not from 0. it's showing it from 180 or nearly 180 degrees of shoulder abduction.
One strange exception I ran into is 43sec into Goniometry Shoulder Abduction & Adduction (by kelly) youtube.com/watch?v=WgTu6HsZZTE it shows adduction measured where 0 is arm up. That seems strange to me, normally  abduction and adduction are measured from the same 0 point.
This video by OTStudentVids shows Horizontal shoulder abduction and horizontal shoulder adduction and takes 0 as out to the side. https://www.youtube.com/watch?v=Zu0WPhfHCWc All makes sense, abduction is one way, and adduction is towards/ across the body.


text:
Is plastoquinone not an electron carrier?
In my book, under the topic chemiosmotic hypothesis, it is given that proton gradient is created by accumulation of protons in the lumen of thylakoids. One of the ways by which the gradient is created is mentioned as below....
' As electrons move through the photosystems protons are transported across the membrane. This happens because the the primary acceptor of electrons which is located towards the outer side of the membrane transfers its electron NOT TO AN ELECTRON CARRIER BUT TO AN H CARRIER. Hence this molecule removes proton from the stroma while transporting an electron.
I suppose they are calling plastoquinone as the H carrier, but then, why is it not an electron carrier? It is involved in electron transport isn't?

By 'H carrier', I think the author means 'a proton plus an electron carrier' or a H atom carrier.
When PQ accepts an electron, is may also pick up a proton from the outside (it now has accepted a H, that is a proton plus an electron).
If the electron is donated to another acceptor, a proton may be lost on the inside. PQ is now back in its original state, but the net result is that a proton has been 'transported' across the membrane (but it is almost certainly not the same proton) and an electron has been 'carried' from donor to acceptor.
(Oxidation is the loss of an electron or electrons. Reduction is the gain of an electron or electrons. But accepting or losing a proton is neither an oxidation nor a reduction, but an ionization)


text:
What is the nature of false negatives on Covid-19 Lateral Flow Tests?
Mainly the thing I'm trying to understand is "how do repeated LFT testing and PCR testing interact with false negatives".
But I think the answer to that is a fairly direct function of what causes false -ves on LFTs.

Is it simply that whatever the testing "pallet" does has a random noise component to it, and sometime you get unlucky?
Is it that the sampling process is noisy, and thus sometimes you get unlucky with the swab and don't catch any viral particles?
Is it that sometimes the body is compromised but immune system controls the viruses presence in such a way that there isn't anything in the upper-respiratory tract?
Is it that some mutations of covid don't trigger the tests?
Something else?

Respectively for the above possibilities, they would mean that:

If you get a false -ve, you could just put new drops form the same sample in another pallet and immediately get a true +ve.
If you get a false -ve, you could take another swab and immediately get a true +ve.
Either:

If you get a false -ve, then that might indicate that LFTs are mostly worthless for you personally. (If your personal immune system fundamentally prevents them doing their job)
If you get a false -ve, you would consistently get false -ves for at least another <x days/hours>

If you get a false -ve, you will continue to get false -ves for the entire course of this infection ... but LFTs might work again on a different infection.
???

And likewise for the interaction with PCR testing:

LFT result has no interaction whatsoever with PCR.
If you had used that particular sample for the PCR test then you would have gotten another False -ve.
If you take 2 samples for LFT and PCR at the same time then you'll get False -ves on both.
LFT result has no interaction whatsoever with PCR.
???

Obviously, plenty of those guesses may not be biologically or statistically possible ... but hopefully they're useful as an indication of the sorts of distinctions I'm asking about and the effects that the mechanics of a False -ve could conceivably have.

Another way to phrase this could be ... "If I take lots of LFTs one-after-another, after a high-risk exposure, and they're all -ve ... does that change the odds of whether I'm actually covid-free?". "What if I take them over the course of a day/week?"

Or "If I tested continuously LFT -ve for 2 weeks, but consistently PCR +ve in the middle ... does that tell me anything interesting/useful about future LFTs"

Is it simply that whatever the testing "pallet" does has a random noise component to it, and sometime you get unlucky?
Is it that the sampling process is noisy, and thus sometimes you get unlucky with the swab and don't catch any viral particles?
Is it that sometimes the body is compromised but immune system controls the viruses presence in such a way that there isn't anything in the upper-respiratory tract?
Is it that some mutations of covid don't trigger the tests?
Something else?

Yes -all of the above, apart from the "immune system controls viral presence to not upper respiratory tract" point. That bit is almost entirely down to viral factors and attachment factors such as the presence of the receptor proteins/sialic acids etc.
Basically the way that lateral flow tests work is that they solubilize the sample, which is then dropped into the well. From there the liquid moves along into the flow part of the test, filtering out large junk and binding to labeled antibodies. When the sample reaches the test line, a second antibody binds to the label on the first antibody, which results in a coloured reaction between two chemical components. The control line does similar, but is designed to detect any antibody or perhaps something that is spiked (added) into the kit as a control. There's a description and video of how these work here.
If any one of these steps goes wrong, from getting a sample without enough virus, to no reaction at the control line (or at the test line), then you could have a false negative. A common problem is not sampling enough virus to work. I don't know how much virus is needed to be added, but in my tests on early versions of these with purified virus (not nasal swabs), I could get down to about 1000 viral particles per test based on qRT-PCR assays of viral concentration. The sensitivity of these tests will depend on the brand and exactly what they are testing for, and should come on an information sheet which you can either get with the test or look up on your health authorities' authorization of the tests.
However, the most common false negative I suspect will be that the test subject has been tested outside the test window, and there is not enough virus present to trigger the test as positive. Generally you need to be between about 3-days and 2-weeks post-infection for these tests to detect SARS-CoV-2 in patient samples.
On the other hand, PCR tests are much more sensitive, and can detect the virus before you are infectious, and long after the infectious window has passed.
There's a good graph of this from a paper by Dankova et al at the International Journal of Environmental Research and Public Health1:

In this case the lateral flow test is marked as "Ag" (for antigen; these tests are often called rapid antigen tests (RATs) too). As you can see the Ag section is quite short and starts well after the infectious period starts.
Now for your next bits:

Respectively for the above possibilities, they would mean that:

If you get a false -ve, you could just put new drops form the same sample in another pallet and immediately get a true +ve.

Possibly, but this would be a failure of the first test rather than a testing/sampling problem

If you get a false -ve, you could take another swab and immediately get a true +ve.

Maybe, this could be a sampling error, nothing to do with the test itself, or a test failure.

Either:
* If you get a false -ve, then that might indicate that LFTs are mostly worthless for you personally. (If your personal immune system fundamentally prevents them doing their job)

No, they shouldn't be personally worthless to you, they don't have anything to do with your immune system or any part of your body. When there is enough virus there, they should detect it.

  * If you get a false -ve, you would consistently get false -ves for at least another <x days/hours>

Maybe - this could be that you've sampled outside the window for the test to work, as you sort of hint at, or that you didn't sample properly

If you get a false -ve, you will continue to get false -ves for the entire course of this infection ... but LFTs might work again on a different infection.

No, this is very unlikely unless you have a dramatic genetic change in the virus such that the antigen is no longer picked up by the test.

And likewise for the interaction with PCR testing:
*LFT result has no interaction whatsoever with PCR.

They don't interact - they don't detect the same thing. The PCR detects the viral RNA, the LFA/RAT detects viral proteins.

If you had used that particular sample for the PCR test then you would have gotten another False -ve.

Maybe, PCR is much much more sensitive and capable of detecting the virus for longer, so a negative on a LFA/RAT doesn't mean a PCR would be negative. False negatives by the PCR tests are very rare as the tests include internal controls to ensure a correct sample has been taken.

If you take 2 samples for LFT and PCR at the same time then you'll get False -ves on both.

Unlikely. Same as immediately above. Repeat sampling of the same sites could remove the necessary material for the test to work. However, generally the PCR swab is taken in the deep nasal tissues, while the RAT/LFA is from shallow and/or throat. The PCR should also work on the RAT/LFA samples, but I believe is less reliable due to the presence of nasal mucus which interferes with the reaction. I'd have to really dig through the literature to find a reference for that, but I think I read one a couple of years ago.

LFT result has no interaction whatsoever with PCR.

Same as 3 points above.
1: Dankova, Z.; Novakova, E.; Skerenova, M.; Holubekova, V.; Lucansky, V.; Dvorska, D.; Brany, D.; Kolkova, Z.; Strnadel, J.; Mersakova, S.; Janikova, K.; Samec, M.; Pokusa, M.; Petras, M.; Sarlinova, M.; Kasubova, I.; Loderer, D.; Sadlonova, V.; Kompanikova, J.; Kotlebova, N.; Kompanikova, A.; Hrnciarova, M.; Stanclova, A.; Antosova, M.; Dzian, A.; Nosal, V.; Kocan, I.; Murgas, D.; Krkoska, D.; Calkovska, A.; Halasova, E. Comparison of SARS-CoV-2 Detection by Rapid Antigen and by Three Commercial RT-qPCR Tests: A Study from Martin University Hospital in Slovakia. Int. J. Environ. Res. Public Health 2021, 18, 7037. https://doi.org/10.3390/ijerph18137037


text:
Do wild birds eat the egg shells of their own offspring?
Background:
I'm a volunteer who is monitoring nest boxes that were established for American Kestrels (Falco sparverius). This spring I've cleaned several boxes that were newly-installed the previous spring, with no intervening monitoring until I showed up a year later. They were obviously used last season (poop, pellets, prey remains), but no egg shells.  So either the inhabitants never laids eggs, or they laid eggs and ate the shells.  Seems to me that the shells would be a good calcium source.
Question:
Do wild birds eat their offspring's egg shell?

Did you try Googling this first? I immediately came up with an answer.

This may unusual for a Q&A site, but we require evidence of an effort on the asker's part that they tried googling (or a book/whatever) to try to answer their own question. Yes/no answers here will be deleted, and this does fall into that kind of answer. The concept is that the more effort you put into your question, the more effort we put into our answers, even to the point of often/usually giving you "proof" that you can trust the answer you receive on this site. You can edit your question by clicking on the edit button. Thanks. (And what a great volunteer "job"! I'd like that.)


text:
Why do some amino acids have more synonymous codons than others?

In the standard genetic code (above), tryptophan and methionine are coded for by a single codon, other amino acids such as phenylalanine by two codons, isoleucine by three codons, and some by four or even six codons.

Is there any explanation for this difference in number?

One possibility that occurred to me is whether, amino acids which are more crucial to a protein’s structure tend to have more synonymous codons? For example, Tryptophan is coded by a single codon, TGG, whereas Proline is coded for by the four codons, CCA, CCG, CCC, and CCT. A single mutation would always change tryptophan, but would have a 1 in 3 chance of leaving proline the same. Is this because a change in proline would be more likely to have a deleterious affect on a protein than a change in tryptophan, and for this reason there has been evolutionary selection to provide proline with more codons?
N.B. This is a different question from the previous general one on the evolution of the genetic code.

Welcome to SE Biology. We do expect people to do a little research before posting to try to answer questions themselves. Theoretical questions like this can only be approached by logical arguments, which anyone with basic biochemical knowledge can try. So ask yourself, would a change in methionine be unimportant? If you consider the mechanism of initiation of protein synthesis, I think you can reject that hypothesis. The fact that this hypothesis was not considered in the question you link to is also suggestive. A simpler idea is that the structurally simpler amino acids evolved first.

@David Yes, you are right that its status as a start codon makes ATG a bad example. I will change it to Tryptophan.

Could you clarify what you mean by “this would make sense”? Things only make sense in relation to a conceptual framework, and in this case you are presumably making some assumption about the way the genetic code evolved. I can guess what your assumption is, and if my guess is correct, almost certainly do not share it, but if you must ask questions like this and want an intelligent discussion you must spell your assumptions out. And changing met to trp doesn’t alter the fact that the situation with met is inconsistent with your proposition. How do you accommodate it?

@David I think I understand now that the unit that natural selection is operating on here is the boundaries between amino acids in codon space, rather than the codons themselves. It is different enough conceptually that I maybe should write another question.

Consideration of the ‘importance’ argument
In some cases it is difficult to say whether changing one amino acid would have a more deleterious general effect than changing another, but one can put forward several examples in which the number of synonymous codons for different amino acids does not support any relationship to structural or functional importance in proteins.
• The fact that there is only one codon for methionine, which is of paramount ‘importance’ as it is essential for the initiation of protein synthesis.
• The difficulty in arguing that arginine (6 codons) is three times more ‘important’ than lysine (2 codons), when these amino acids are equally abundant in E.coli and lysine is involved in the catalytic function of numerous different types of enzyme.
• The difficulty in arguing that the non-catalytic leucine is ‘important’ enough to have 6 codons, when there are another 3 for the related isoleucine and a swathe of other amino acids with aliphatic side-chains, mutation to which may be allowable.
• The fact that unique indole ring of tryptophan (1 codon), as well as providing a large aromatic side-chain,  plays an essential catalytic role in a variety of enzymes such as oxidases.
Is selective pressure to minimize amino acid changes likely to have been important in the evolution of the genetic code?
The genetic code today is the same in almost all organisms, and the variations in the exceptional species are only minor. In the generally small and isolated mitochondrial genomes the selective pressure for variation would seem to have been to reduce the number of tRNAs needed to decode the mRNA. This is consistent with the “frozen accident” idea (originally proposed by Crick) — once the genetic code was to a certain stage it become frozen because changes would have been lethal. Remember changing the assignment in one codon would affect all proteins using that codon, in contrast to a mutation of a single codon in a single protein to a different amino acid. Further, any codon reassignment would affect the anticodon of the tRNA corresponding to that changed codon — often interacting with more than one codon and dependent on recognition by the amino acyl-tRNA synthetase for attachment of the correct amino acid.
An alternative explanation for the conundrum
Disclaimers:
We clearly cannot know how the genetic code evolved, so all we can hope for is to find the explanation with the fewest flaws in relation to scientific knowledge at that time. Some of this answer appears to have come subconsciously from the 1968 article by Crick on the Origin of the Genetic Code, which I re-discovered after having formulated it. Other aspects (no doubt the weaker ones) are my own, but must surely have been considered by others in the 60 odd years since the elucidation of the genetic code.
Summary of answer:
My suggestion for the difference in number of synonymous codons is that many of the amino acids with only two synonymous codons — and those with only a single codon — may have evolved later than those with three, four or six synonymous codons.
Answer
It is not unreasonable to suggest that the original genetic code specified fewer than the 20† contemporary encoded amino acids and the termination signal. This would have been because of the primitive stage of evolution of metabolic pathways for amino acid synthesis (presumably catalysed by ribozymes). There may even have been fewer than 16 amino acids, although with a triplet (rather than a duplet) codon length because of geometric and energetic considerations relating to the interaction with tRNAs (themselves ignoring or interacting promiscuously with the third codon position).
So the 16 blocks of codons would have been assigned to those initial amino acids, with some perhaps taking over two blocks (Leu?). As metabolism developed, new amino acids would be added, either occupying ‘empty’ blocks or taking over half of an existing block (Phe in the Leu block?). As new amino acids were added with more functional possibilities then simple aliphatics or aromatics, and more sophisticated proteins arose, it would have become increasingly difficult to change the genetic code because of the disruptive effect this would have had.
Any theory should clearly consider, as the OP did, the amino acid tryptophan with its single codon — but, likewise, also methionine. One can envisage a situation in which the code had developed to encompass the other 18 amino acids and perhaps four stop codons, but without a specific initiation codon. Others have argued that methionine emerged late as the initiating amino acid (an early role as another aliphatic is at odds with the high metabolic cost of its synthesis); suffice it to say here that Met is chemically similar to Ile, which one assumes it displaced in the AUN block, and such a change would therefore have been generally tolerated by proteins.
I think it likely that, because of its complex structure, Trp was added late (last?) and replaced a termination codon. This is because one can argue that it would have been less disruptive than replacing an amino acid codon. First, termination does not involve tRNA and aminoacyl tRNA synthetases — the codons are recognized by release (termination) factors. This would only require mutations in these factors, which show differences between eukaryotes and eubacteria. Second, the effect of a mutation in a stop codon is just a longer C-terminal segment to the protein: termination still occurs when one of the remaining stop codons is encountered by chance. Variation in the C-terminal segments in proteins are widespread between species, in different isoforms of a protein within a species, and in the strategies viruses use to expand their repertoire of proteins from a compact genome.
†The additional amino acids selenocysteine and pyrrolysine, although of interest, are not relevant to this argument.

That is a good point, but I am not sure that, at least before all 64 codons were assigned, all codons were frozen. With an unused codon available the genetic code can change by 1. assigning an amino acid to the codon, 2. switching all proteins to the new codon, then 3. assigning something else to the newly free codon. The fact that the code can't be changed without destroying almost every protein doesn't come in to play until the codons are all being used. That implies that natural selection may have influenced the choice of many codons, even if as you point out it can't change them today.

Having fewer codons makes an amino acid less likely to be mutated to, while having more codons makes an amino acid less likely to be mutated from. If a particular amino acid would drastically change protein expression if it either appeared or disappeared (for example fMet, the start codon in bacteria), it is not clear whether natural selection would prefer to guard against its appearance (by giving it fewer codons) or disappearance (by giving it more). If natural selection does determine the genetic code, it would have to do so by acting in a more subtle way that could differentiate between those two cases.

Your answer assumes that the consequences of mutation were key factors in determining codon assignments during evolution. I very much doubt this. The whole question is complex, but has been discussed repeatedly since the genetic code was deciphered. Before expressing an opinion on this it is wise to consider such previous arguments. I regret not having had time to do so, which is why I have been reluctant to get involved. However, I'll post an "answer" some time next week  explaining why I think your assumptions — and hence your answer — are invalid.

This answer ignores a whole bunch of interesting biology such as codon bias in different organisms etc. It is an interesting idea you have had, but you should back it up with some references that show this is actually the case.

@bob1 — Could you explain what codon bias in different organisms has to do with the evolution of the genetic code?

@David not off the top of my head, but I think it could play into the evolution somehow - energetic costs? availability of substrates for generation of bases?  Could simply be a founder effect.


text:
Why do we have to scrape the inside of the cheek twice to get the sample of epithelial cells?
I was studying the theory of a practical exercise of preparing temporary slide of animal tissue from my laboratory manual.
In the section about epithelial tissue, the first few steps of the procedure contains the following.

• Rinse your mouth well with water.
• Gently scrape the inside of your cheek with the broad end of a clean
toothpick. Discard this material.
• Scrape again, and spread these cells gently on a clean slide. Add a drop
of 0.9% NaCl solution or physiological saline and a drop of methylene
blue with the help of a dropper.

Why is it mentioned to scrape and discard the first time and scrape again?
I've tried searching about it on the web but couldn't find any mention of scraping twice to obtain the sample, nor it is mentioned in my book. There doesn't seem to be any apparent reason to scrape twice to me.

What do you think? What have you found in your research? The Biology.SE community has agreed that questions that show little or no prior research effort are off-topic on this site unless you have shown your attempt at an answer. Please [edit] your question and tell us where you've looked for answers, what you do know about the topic, and where exactly you still have questions. Unresearched questions may be subject to down-voting and closure. Please see our homework policy for more information.

@MattDMo I've added info. accordingly.


text:
Aminoacyl-tRNA synthetase and the degeneracy of the genetic code
Aminoacyl-tRNA synthetase catalyses the covalent linkage of amino acids to tRNA.  The aminoacylated tRNA is then bound to mRNA on the ribosome in a codon–anticodon hydrogen bonded interaction.
In order for several different codons to correspond to the same amino acid — according to the degeneracy of the genetic code — one or more of the following must be true:

Each amino acid is recognized by several different aminoacyl-tRNA  synthases, each one able to aminoacylate a tRNA specific for a different codon for that amino acid.

One aminoacyl-tRNA synthetase can aminocylate the several different tRNAs for a specific amino acid.

A single tRNA for a particular amino acid can bind to several different mRNA codons.

Which of these is true?

This topic is dealt with in most biochemical and molecular biological texts — there is a useful summary in Alberts et al.. However it may be useful to clarify this point as this is a fundamental question, the answer to which could not be anticipated.

Both 2 and 3 are correct, but 1 is incorrect.

As regards 2, it turns out that there is a single aminoacyl-tRNA synthetase for each amino acid and this recognizes all the different tRNAs for that amino acid. This is even true of initiator and elongator tRNAs for methionine. This implies that even though the anticodons of these tRNAs differ (or their conformations in the case of the two tRNAs for methionine) there is some common distinguishing feature of the tRNAs that the enzyme recognizes.
As regards 3, this relates to the phenomenon of wobble, the possibility of specific relaxed patterns of hydrogen bonding at the third position of the codon. This decreases the number of tRNAs required to decode the codons corresponding to a particular amino acid.
In principal neither of these features is neccesary for the degeneracy of the genetic code. They just reduce the number of molecular species (enzymes and tRNAs) required for decoding. One assumes that the practical advantage of this was sufficient to ensure its development and retention.
Coda
Nature is an anarchist, and although the above statements are generally true there are always exceptions. For example, some bacteria have developed ways of managing with fewer than the 20 expected aminoacyl-tRNA synthetases. But if one starts by understanding the general situation one is then in a position to accommodate the exceptions. A highly-cited review of aminoacyl-tRNA synthetases can be found in Ann. Rev. Biochem. for 2000.


text:
Do microorganisms have complex social behaviors like dominance hierarchies, just as "higher animals" do?
I used to believe that dominance hierarchy (a.k.a. pecking order) exists in higher order animals and is associated with complex social behavior. See the wikipedia definition of dominance hierarchy and pecking order which is restricted to animals. For example, a pecking order can be an order decided by the brightness of feather, without straight competition or direct fight.
However, I recently came across some examples on the complex social behavior of microorganisms, dominance hierarchy of microorganisms and bacteria, dominance hierarchy and hierarchy of exploiting in bacteria, and pecking order of pastures and plants.
For  a specific example, here is a comment of a MIT paper published on Science about "social structure of microoganisms":

They reasoned social structure can reduce conflict within populations of plants and animals and determine aggression towards competing biological populations. "Think of a population of lions in the Serengeti or a population of fish in a lake," said Cordero. But could the same be true for populations of bacteria?

The answer to this question seems to be positive, according to the abstract of the Science paper:

In animals and plants, social structure can reduce conflict within populations and bias aggression toward competing populations; however, for bacteria in the wild it remains unknown whether such population-level organization exists. Here, we show that environmental bacteria are organized into socially cohesive units in which antagonism occurs between rather than within ecologically defined populations.

So I am very confused.

A better question than last time. Note that just because they used the words "dominance" and "hierarchy" in them, doesn't mean that the papers are talking about a social order. The first plant link is about genetic dominance/recession in alleles within the plant. Can I suggest you investigate the terms mutualism and antagonism in ecology?

@bob1 You are right that those two things are only distantly related. I will delete it.

Similar to my comment on your other similar question and also bob's...have you read the papers you cite for "dominance hierarchy"? What concept are they talking about? Does it relate to what you know about "pecking order" in social mammals? I think they are each clear about what they mean; the same words can be used in different contexts to mean different things.

Short answer
All of these papers describe competition; that's all they share in common. Competition is ubiquitous in nature; it need not even involve organisms that have ever encountered each other: when you put two organisms together that use some of the same resources, competition occurs. Just because competition occurs does not mean "complex social dominance hierarchies" occur. Just because someone talks about a "hierarchy" it does not mean that hierarchy is the result of a social agreement for weaker individuals to defer to stronger ones to avoid conflict; "hierarchy" just means "we are putting these entities in an ordered list"; the order and meaning of that list depends entirely on the conditions and criteria used to arrange that list. When someone says "hierarchy" or "order" you have to read what they mean by hierarchy/order and not assume that it is related to any other hierarchy you have read about before. If you have some sports teams, you could rank them according to their win/loss record, according to an Elo-based ranking system, according to how much you like their team colors, according to a weighted average of expert/sportswriter opinions about how good the teams are, etc. The teams don't even have to meet each other for most of these measures, they certainly don't need to agree among each other about who is stronger for you to be able to put them in a hierarchy according to your preferred criteria, and even if you think you've designed the best hierarchy possible, their pairwise interactions may not be what your hierarchy predicted.
Long answer
The type of dominance hierarchy described on Wikipedia is an evolutionarily stable strategy among social animals. In short, by establishing a hierarchy, fighting is limited. Fighting is costly and dangerous: wounds that may seem minor can result in deadly infections, and any effort towards a fight costs energy/calories.
Your question cites several other situations where authors have used "dominance hierarchy" or "pecking order" to describe the results of competition, but not all competition needs to have this social deference-to-established-strength aspect.
With respect to the specific papers you talk about:
https://royalsocietypublishing.org/doi/epdf/10.1098/rspb.2003.2338
This paper is about competition among yeasts. It does involve interference competition and exploitation counter-strategy. However, there is no complex social hierarchy, this is all competition happening on a molecular level. Species/strains that produce and are resistant to a toxin get the benefits of the toxin but also carry the costs of producing it. The authors study a system where growth patches are isolated and from each other and strains spread from patch to patch. When present in the same patch, the toxic strains can often defeat the sensitive strain. However, sensitive strains can be successful in places where the toxin producers are absent, and since they don't need to bother to produce toxins, they grow and spread faster. Depending on the parameters of the environment, the increased growth ability comes into an equilibrium with toxin production.
Nothing social needs to happen for this to occur, none of the yeast strains in these models are behaving in any different way, it's merely a result of the cost-benefit of producing a toxin.
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC106711/
There is absolutely no intraspecies social competition in this paper. They have squid and bacteria that have a symbiotic relationship. If you take the bacteria from one species of squid, it can survive in other squid species, but it doesn't do as well as the bacteria native to that squid, and vice-versa. The closer the squid are phylogenetically, the closer their bacteria are, and the more similar cross-collaborations are better than dissimilar ones. The "hierarchy" here is completely environment-dependent; from the perspective of the bacteria, you have a different hierarchy for each squid species.
https://onlinelibrary.wiley.com/doi/full/10.1046/j.1420-9101.2003.00545.x?sid=nlm%3Apubmed
In this paper, they study a slime mold that produces a fruiting body to spread. The fruiting body consists of a stalk and spores; in their terms, they call "dominant" strains the ones that produce more of the spores. However, the stalk is important for effective reproduction. Therefore, strains that produce more spores are "cheating" a bit, and this strategy is only effective when other strains are present to make up the stalk. This is a complex interaction, but doesn't rely on any social hierarchy; the "hierarchy" here is simply the ratio of spores to stalk produced by each strain, in other words, the level of "cheating". Being "high" on this hierarchy aka a "cheater" is only good when there are lots of non-cheaters around.
https://www.jstor.org/stable/3565800
Here they are growing plants together in a plot. They rank these plants according to how they grow in competition with other plants. There is no social interaction here at all. The paper shows how hierarchies can be non-linear due to different demands for resources that appear when different species compete. Therefore, Plant A might outgrow Plant B, and Plant B outgrows Plant C, yet Plant C outgrows Plant A if each are isolated in a bi-culture. Real ecosystems contain more varied conditions than an experimental plot, so in the real world you'll find different niches where each plant exists. Despite using the term "dominance hierarchy" these authors aren't talking about anything but relative abundances in different co-culture scenarios.
The last link is to a book on plants; I won't address it directly here, because it's unclear what part of the book is being referenced, but I am guessing it's describing something similar to the Aarssen article.

Ok very detailed answer. Much appreciated. I get your point, especially your interpretation on those articles. So are you suggesting that microorganisms do not have complex social behaviors?

@HighGPA "Complex" is... often a meaningless word, unless you specify ahead of time what you mean by "complexity" and how to measure it. Also, no matter how you define/measure it, it's usually used in relative terms. The same is true of the word "social", as you can use it to refer to the types of relationships that form among humans, or use it to mean generally "interactions between two or more organisms of the same species".

I also agree with you that fighting in costly. However, I would like to point out that your "complex social hierarchy" is also determined by competition, for example, that peacocks competing the brightness and largeness of their feather. Such feather is also very costly to grow. So I wonder if there is a clear cut between competitive hierarchy and dominance hierarchy.

@HighGPA The peacock example is better defined by sexual selection than "pecking order" or a dominance hierarchy; I don't believe they participate in those social structures.

Well I thought the alpha peacock get more sexual and material resources?

After reading your answer, here is my understanding: the cited articles are mostly about direct competition, and the order determined by direct competition is not "dominance hierarchy"; however, if some orders in bacteria is determined by signaling, without any direct competition for resources, then this could be called the "dominance hierarchy". Is my understanding correct?

@HighGPA I believe peahens form harem groups during breeding season; as such, there is not so much an "alpha peacock" and a bunch of subordinate males, but rather a single male peacock and the peahens that choose to mate with him.

For "dominance hierarchy" I think you can apply that label to whatever you want, as some of these papers have, but when doing so you need to explain what you mean. Just because they say "dominance hierarchy", whatever the form, does not mean it comes with all the baggage of "dominance hierarchy" used in another context. I understand this may feel messy and confusing, but...well, welcome to biology - this sort of thing is common. It's incredibly important in understanding biology to know what terms mean as people use them, and not to overgeneralize.

Let us continue this discussion in chat.


text:
Production of “Green Hydrogen” by photosynthesis
In a popular science book from the 1970s I read about experiments to tweak some microorganisms to produce elemental hydrogen by photosynthesis.
Is this a feasible way to produce “Green hydrogen”? What kind of microorganism can be used for this task, and what is the efficiency of that process?

Photosynthesis
The aspects of photosynthesis pertinent to this question are as follows:
The overall chemical transformation in the light reactions of photosynthesis is:

2H2O + hν → 4H+ + 4e– + O2

The hydrogen ions are produced in such a way as to create a gradient that is used to generate ATP, the electrons are used to reduce NADP+ to NADPH. Together these allow the reductive synthesis of carbohydrate sugars from CO2.
This process involves two photosystems — PSII and PSI — shown below, the latter of which is most pertinent to this topic.

Coupling Photosynthesis to Hydrogen Production
The basic objective of modifying photosynthesis to generate hydrogen — biophotoloysis — involves using the electron to reduce the hydrogen ion to hydrogen gas, rather than allowing it to be used to oxidize NADP+.

2H+ + 2e– → H2

So that the overall reaction becomes:

2H2O + hν → 2H2 + O2

(Of course, this will be at the expense of energy and reducing power for carbohydrate synthesis.)
Using Hydrogenase for the Catalysis
The enzyme, hydrogenase, can catalyse the reduction of hydrogen ions shown above. This enzyme is rare in eukaryotes and absent from higher plants. It is thought to be very ancient, and may have originally been involved in energy generation from hydrogen in early evolution. One of the roles it plays in contemporary organisms is in reoxidizing NADH generated during certain fermentations in bacteria such as the Clostridium family — hydrogen is the gas produced in gas gangrene caused by Clostridium perfringens.
Certain photosynthetic organisms — notably the microalga, Chlamydomonas reinhardtii, and the photosynthetic cyanobacteria — also contain a hydrogenase in their chloroplasts. The activity of this is generally low, but appears to be coupled to photosynthesis in certain circumstances. This is through the reduced ferredoxin produced at PSI transferring its electron to the iron or iron–nickel centre of the hydrogenase:

The physiological importance of this coupling is not fully understood but its discovery offered the possibility of using it to produce hydrogen from water.
Commercial Application — Approaches and Problems
The focus of many of the attempts to exploit this coupling to produce hydrogen on a commercial scale have used Chlamydomonas reinhardtii, where is was shown that deprivation of sulphur enhanced the evolution of hydrogen significantly. From a commercial standpoint the price at which a fuel can be sold depends on the cost of its production, and to be viable this has to be competitive with alternatives. Hence it is important to maximize the production of hydrogen, and considerable effort has been directed towards this. However, we are not yet at a stage where photosynthetic hydrogen can compete with other fuels.
The two main factors limiting widespread commercial exploitation of this process have been the sensitivity of the dehydrogenase to inhibition by the oxygen produced in the process, and the low efficiencies of conversion of solar energy (<2%). The former problem is, in fact, one of the reasons for using Chlamydomonas reinhardtii, as it can perform oxidative respiration, lowering the concentration of oxygen. Attempts have been made — and are presumably still being made — to engineer the hydrogenase to reduce its sensitivity to oxygen, to control the oxygen concentration, and to improve the photosynthetic quantum yield by engineering PSII. Increasing the efficiency of photosynthesis itself is a more difficult problem in the eukaryote, Chlamydomonas, and efforts in this respected have mainly been directed at photosynthetic bacteria.
Literature
My knowledge of this field is based purely on a perusal of sources found in a literature search. I was actually surprised to discover that there is at least one journal dedicated to hydrogen as an energy source. Here are the papers I consulted:
Bolatkhan et al. (2019) Int. J. Hydrogen Energy 44, 5799–5811
Oh et al. (2013) Biohydrogen, Ch.3, pp. 45-65 Elsevier.
Ni et al. (2006) Fuel Processing Technology 87, 461–472
Hemschemeier, A. and Happe1, T. (2005) Biochem. Soc. Trans. 33, 39-41


text:
When a dog sniffs up organic material that contains DNA, is it possible his genome incorporates it?
I'm not kidding.  Was just watching my German Shepherd sniffing away at a new Amazon box.
...realized she sniffs vast array of DNA from organic material when on a walk: millions of mammalian, plant, fungi, etc. DNA molecules.
It would promote fitness of a species if DNA has a method of feature exploitation from cross-species DNA "pollination."
Wondering if anyone has heard of research
I was in microbio a short time while inventing the software for first DNA editing robot for Inscripta back in 2017.  Process was CRSPR.  Over 1000 steps.  If a simple bacteria has the capability to remove a DNA exploit from a virus, should not the reverse be possible?
If I was God, I'd dub that in for sure.
Would allow faster local adaptation, should humanity get to other worlds.

What's your question?

Whole DNA cannot travel from the olfactory system (or the digestive system, for that matter) into cells' nuclei and be transcribed/translated. That would be very bad if it did - would you like to start expressing fruit genes every time you smelled a fruit, or bacterial/fungal genes when you smelled dirty shoes? "Adapting to the local environment" does not mean expressing local genes.

S Pr: When a dog sniffs up organic material that contains DNA, is it possible his genome incorporates it?

MattDMo: Good point. But if a dog or a human sniffs a virus, can it get at our cells and reprogram them to replicate the virus?

But that's what viruses do, it's part of the definition. I'm unclear how you might have been working in microbiology and not have encountered the basics.

Animals don't really smell DNA or any nucleic acid
I am not aware and really truly doubt any mammalian (nor vertebrate/invertebrate) organism can detect DNA/RNA via the olfactory system (nose, antenna, sensilla, etc.).
Typically we think of the olfactory system as reserved for volatiles (odors), and the gustatory system for tastants like salt and sugar and bitter compounds. A dog cannot smell DNA as typically present in nature in ecologically-relevant contexts, as DNA is not volatile at all (it's a giant molecule, and things you can smell are typically very small in molecular weight, with a low vapor pressure). Even if present in droplets, it wouldn't be detected by olfactory receptors, which are the interface responsible for transducing chemical "odor information" into electrical information processed by the nervous system. I once also answered a question about whether one can smell or taste viruses and spores; the answer is categorically no. You can only smell or taste some of its components can can become airborne. You could perhaps smell some outcome of a viral infection or the metabolites or by-products of mold, but not the cells or viral particles themselves.
Can DNA be incorporated in the nasal/olfactory epithelium?
Oh, no. It's even less likely is the possibility that DNA is taken up by cells in the olfactory epithelium. And even if it was to be taken up by cells, genome incorporation of random DNA is pretty much impossible. That's extremely unlikely, on top of extremely unlikely.
However, can we taste nucleic acids like DNA?
You can perform this test yourself at home! The easiest way would be to try isolating some DNA from strawberries, which are octaploid and a great source of easy DNA! If you were to taste it, according to one person, you would find it

salty and slimy after sitting in his mouth for as long as he could
stand it. Conclusion: strawberry DNA is best eaten accompanied by the
rest of the strawberry.

Probably salty due to the sodium and potassium ions that neutralize the charge of the phosphate backbone that is present in DNA.


text:
Does anybody know what sort of fossil this could be?
enter image description here
Found in rural North Dakota. Small rock with black circular indentations.

Welcome to the site. Can you please provide more pictures, with better lighting of the "fossils". It may not be a fossil at all, but rather some sort of inclusion in the rock.  Fossil ID is on-topic for Earth Science SE, but you would need to request migration - don't cross-post!

it looks like marks made by animals living on the stone probably made under water,it is very unlikely to be fossils the rock is rounded most likely when submerged under water.

Thanks guys! Any bit of info helps.


text:
Which anaerobic environments would exist without aerobic life?
All of the anaerobic environments I can think of are that way because a layer of aerobic life above them separates them from oxygen. If the aerobic life were removed, the anaerobic compartment would equilibriate with the partial pressures of the atmosphere, however slowly, and it would remain at equilibrium until competition between aerobic cells once again pushed the oxygen boundary to the top.
After the great oxygenation event, there was oxygen in the atmosphere but no layers of aerobic life to stop it from diffusing to the bottom of the ocean or deep underground. If there are anaerobic environments that exist independently of aerobic life, our anaerobic ancestors could have survived there until aerobic respiration or tolerance evolved. Otherwise aerobic tolerance would have had to have been present before the great oxygenation event for anything to survive at all.
Which is closer to the truth? (An environment that takes extremely long to equilibriate with the atmosphere, long enough for evolution to be fast in comparison, would count as isolated for the purposes of this question since it would serve as a reservoir of life from which aerobic respiration could emerge.)

well since most of the oxygen is made by aerobic life, anywhere they want.

A lot of chemicals would have buffered the oxidation event. Especially Fe2+ in water oxidising to Fe3+.

In the question body it says:

After the great oxygenation event, there was oxygen in the atmosphere but no layers of aerobic life to stop it from diffusing to the bottom of the ocean or deep underground.

This statement isn't justified, given that the Great Oxidation "Event" developed over one to two billion years. It took half a billion years for the atmospheric oxygen concentration to go from nil to around 3%.  This is a large amount of time for evolution to select organisms that can tolerate the presence of free oxygen at various levels.  There would be a wide range of environments with differing levels of oxygen, depending on many factors including distance from the oxygen emitters (cyanobacteria) and the nature of the intervening materials.  The atmospheric oxygen concentration remained at fairly stable levels of 2% to 4% for nearly a billion more years, giving more time for evolution of oxygen-tolerant and -consuming organisms to evolve. Thus, to the extent that anaerobic organisms depend on aerobic organisms to shield them from oxygen diffusion, there would have apparently been plenty of evolutionary time for that to occur.

Deep sea hydrothermal vents release reducing chemicals like sulfide ($S_2^-$) and ferrous iron ($Fe^{2+}$). These chemicals establish anaerobic conditions abiotically[1].
[1] "M. jannaschii uses sulfide (S2-) for growth and energy, which is good because sulfide is present in high levels in the vent fluid. It reacts with oxygen to establish anaerobic growth conditions for the archaea."