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Random Mutagenesis vs Directed Evolution as Strategies to boost expression
Do people use random mutagenesis (say using UV) to generate host variants that have high expression of a metabolite / enzyme? I've seen it mentioned as a strategy but it confuses me as to why.
How does that compare to using Directed Evolution for the same purpose? Intuitively, random mutagenesis seems highly inefficient (since you have no control over which genes you mutate) given the combinatorial explosion even in something like E Coli's genome size. Am I making a mistake in this reasoning?
What are situations where one may prefer random mutagenesis over directed evolution? Are there any pros and cons to consider?

Well, you already mentioned it. Random mutagenesis (with UV or chemical mutagens) targets the entire genome and you select the organism for certain traits.
In a directed evolution experiment, you are just targeting a gene or a group of genes. With the progress of synthetic biology, it might be possible to synthesize small genomes (already shown by Craig Venter and group) and apply directed evolution methodology genome wide. This would still not be considered directed, as the mutations are actually random over the genome.
The pros and cons depend on what exactly you aim to do.

Thanks! The aim is something like this: I'm interested in getting yeast to express  a secondary metabolite by inserting a hetrologous gene. The goal is to develop & select a yeast strain that produces high levels of the precursor substrate.

@curious_cat That is the ultimate goal. In that case you can go for random mutagenesis. However, if you are improving the catalysis rate of a specific enzyme, lets say, then apply directed evolution to that specific gene.

Thanks! So it isn't entirely impractical to attempt random mutagenesis? i.e. With 12 Million base pairs & random mutation is it a viable strategy? Have other attempts using similar approaches succeeded?


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Why is DNA Shuffling more efficient than Point Mutation?
In a related post on Biology-SE the following insightful comment was made:

The advantage of DNA shuffling over introducing single mutations is
  that you have to screen fewer mutants and the activity/stability of
  the protein could be improved several hundred fold more.

Is there a mathematical argument as to why DNA Shuffling should be more efficient than introducing point mutations?
Even supposing each DNA Shuffling / Point Mutation step is equally "expensive"  (and in reality DNA shuffling seems much more work) they are both generating one variant per-step, right? So why is the variant in one case grossly better than the other?
Related Post: Directed evolution: Point mutation vs Insertion-Deletion vs Shuffling

You should read this paper. Here is the gist of what you are interested in:

Because most point mutations are deleterious or neutral, the random point mutation rate must be low and the accumulation of beneficial mutations and the evolution of a desired function is relatively slow in such experiments. For example, the evolution of a fucosidase from a galactosidase required five rounds of shuffling and screening before a >10-fold improvement in activity was detected4. Naturally occurring homologous sequences are pre-enriched for 'functional diversity' because deleterious variants have been selected against over billions of years of evolution...
Although shuffling of a single gene creates a library of genes that differ by only a few point mutations1, 2, 3, 4, 5, 6, the block-exchange nature of family shuffling creates chimaeras that differ in many positions. For example, in previous work a single beta-lactamase gene was shuffled for three cycles, yielding only four amino-acid mutations3, whereas a single cycle of family shuffling of the four cephalosporinases resulted in a mutant enzyme which differs by 102 amino acids from the Citrobacter enzyme, by 142 amino acids from the Enterobacter enzyme, by 181 amino acid from the Klebsiella enzyme and by 196 amino acids from the Yersinia enzyme. The increased sequence diversity of the library members obtained by family shuffling results in a 'sparse sampling' of a much greater portion of sequence space15, the theoretical collection of all possible sequences of equal length, ordered by similarity (Fig. 4). Selection from 'sparse libraries' allows rapid identification of the most promising areas within an extended sequence landscape (a multidimensional graph of sequence space versus function)

DNA shuffling would be more efficient if you already have a big repertoire of variants (which have a lot of point differences among them; see Cohen, 2001). When you do not have that, then introducing point mutations using error prone PCR would be better.
A detailed mathematical framework to compare these two methods, is not available. However there are many mutation models and there is at least one model for DNA shuffling too (Sun F., 1999).
Basically, for a point mutation experiment the number of mutants will depend on the error rate of polymerase. In a DNA shuffling experiment you are just creating random combination from existing variants. So, which one is more efficient depends on these (and many more) parameters. Moreover, the "efficiency" also depends on what experiment you are trying.

source: Stack Exchange

license: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/3.0/

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Why cold water results in cough
I have a pretty simple question which always kept me intriguing since childhood and I couldn't find its precise answer yet. Why do we cough by cold water. Recently I had flu and found very strange correlation between cold water and coughing. Even seconds after drinking cold[er] water, I started coughing as you all would have experienced as well. So why does it happen?

@rg255 I think the answer below addresses the question and I don't find any reasons why its opinion based if there is a clear linkage with tract called "Respiratory Mucosa". Rest, you better know than me

I am no expert in this but I had asked my doctor the same question the last time I fell sick and I'll try to paraphrase his explanation.
"There is this protective lining on our respiratory tract called "Respiratory Mucosa" that acts as the first line of defense against pathogens. But when you consume cold food and beverages , this membrane gets congested or shrinks such that it exposes the respiratory tract to pathogens . This is more prevalent if your body has an elevated temperature such as after spending a lot time outdoors during summer or after a workout. Cold food as such doesn't make you sick but they make it easier for the pathogens."
A quick Google search turned up this
http://www.thehealthsite.com/diseases-conditions/why-does-drinking-chilled-beverages-cause-sore-throat/#
Your cough could be a symptom of having a sore throat from infection.

any source to add?

Reference added.

Can you add any authoritative reference, like a journal or a well cited expert's blog/website? As for thehealthsite.com it is absolutely uncited and there's no mentioning who the author is.

all_licenses: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/3.0/


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What is the reason for 'microcytic' anaemia?
I know the causes for microcytic anaemia are Fe deficiency, prolonged inflammation, Thalessemia, Sideroblastic etc. All these logically point to a decrease in Haemoglobin synthesis - either Haeme or globin.
But my doubt is, what exactly causes the microcytosis? Lesser Hb can explain the hypochromicity, but how does it explain the decreased MCV?

The reduced iron availability affects the red cell size during differentiation.

The presence of red cells with reduced mean corpuscular volume (MCV) reflects reduced hemoglobin synthesis, which can have several causes. It can result from defects either in iron acquisition or availability, or in heme or globin synthesis. Experimental data in mouse demonstrated that early mitotic divisions are accompanied with no reduction of MCV, whereas during differentiation mitotic events are associated with a substantial reduction of MCV[1]

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2649346/

Coopersmith A, Ingram M. Red cell volumes and erythropoiesis. II. Age: density: volume relationships of macrocytes. Am J Physiol. 1969;216:473–82.


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What makes something food?
From my (limited) understanding, animals get energy from food by breaking chemical bonds between molecules.
There's a lot of water here [citation needed], so it seems like natural selection would favor any organism that could live off it. Why, in the history of life on earth, hasn't an organism appeared that eats water?
Same applies to dirt, rocks -- (although maybe lichen eat rocks? I'm fuzzy on lichen.)
Or air?
Why are humans and other animals like us only equipped to eat certain types of molecules, when others are so much more plentiful?

Because breaking H2O into H2 and O2 is an endothermic reaction: that is, it takes energy to break the bonds.  Food has molecules that can be broken down, or combined with other molecules such as oxygen, in exothermic (that is, energy producing) reactions.

Food has two purposes: providing energy and providing raw material for growth. Water can only be broken down into hydrogen and oxygen which requires energy input, and water isn't by itself a raw material for anything other than ice. It is however important to life on Earth as a solvent (non-polar and electrolytic), and component of some structural chemicals (hydrates).

You don't get energy from breaking chemical bonds, you only get energy from making chemical bonds, while breaking chemical bonds requires the input of energy.  However, in practice chemical bonds are always broken as others are formed, and the net number of bonds is generally constant.  Otherwise you would end up with free radicals, which are highly reactive and looking to form a chemical bond with the next thing it touches.
For example, the burning of glucose in respiration involves the breaking of C-C, C-H, C-O and O-H bonds in glucose and O=O bonds in oxygen, with the simultaneous formation of C=O bonds in CO2 and O-H bonds in water.  But the C=O bonds in CO2 and O-H bonds in water contain much less energy than the bonds we break within in glucose.  So the key is that the bonds formed contain less energy than the bonds broken, and energy is released overall.
And actually it's a bit more complicated than that, because you also have to consider entropy, which is how disordered the system is.  Generally the more molecules in a system, the more entropy, so breaking a few big molecules down into lots of small molecules tends to release energy by increasing entropy.
So to answer your question, "food" is what we can draw useful energy from in the process of turning it into something else as we make and break chemical bonds.  We can't do this with water, or indeed with rocks, because their chemical bonds would require more energy to break than the energy we'd get back out by forming alternative bonds with anything else.

Oh, that's really clear! So to attempt to parrot it back to you: animals that eat food are turning high-energy bonds into low-energy bonds, and using the energy that results. The low-energy bonds that get created in this process are in things like water and CO2, which get turned into high-energy bonds again by plants (they get the extra energy from sunlight).

So if something is "food" to an animal, it's probably because it has a lot of high-energy bonds.

Yes, that pretty much sums it up.  Ultimately, food stores energy in high energy bonds, and we extract the energy by breaking them and replacing them with low energy bonds in CO2 and water.  In fact most organic compounds contain high energy bonds, which is why they burn in air to give out energy (the heat and light of a flame), although we haven't evolved the enzymes to use them all as food.

So we're talking about bonds that have a lot of potential energy. Are these always bonds that are easy to break, or do the two categories just have some overlap but are unrelated?

The energy of a bond and whether it is easy to break are not directly linked as it depends on the energy of the transition state, but generally the more energy stored in the bond, the easier it is to break and the less energy required to break it via the transition state.  However most enzymes work by reducing the energy of the transition state so that bonds break and form more easily, and thus they speed up the reaction.

PS. A good example to illustrate this, is that gasoline does not spontaneosly combust in air, you need an initial spark to pass through the transition state, then the energy released allows the rest of the reaction to proceed via the transition state.  Similarly, glucose does not spontaneously combust in air and you need enzymes to reduce the energy of the transition state so that the reaction goes at body temperature.

Plants use both water and air as food using photosynthesis where water is split and recombined with carbon dioxide from air to make glucose.

Overall, the chemical reaction of photosynthesis is as follows:

Light energy + plant enzymes
6CO2 + 12H2O ------------------------------------------------> C6H12O6 + 6O2 + 6H2O
...which means that it takes
six molecules of carbon dioxide plus
12 molecules of water

in the presence of light and the proper enzymes in the cell, to make
one molecule of glucose
6 molecules of oxygen
6 molecules of water

Energy is released when glucose is burnt

about 16 enzymatic rxns
  C6H12O6 + 6O2 + 6H2O ------------------------------------> 6CO2 + 12H2O + ENERGY

...which means that
one molecule of glucose in the presence of
six molecules of oxygen and
six molecules of water

can be "burned" to release stored energy as well as the "waste" products of
6 molecules of carbon dioxide and
12 molecules of water

http://www.bio.miami.edu/dana/dox/photosynthesis.html

"Plants use both water and air as food using photosynthesis ... to make glucose." - No, glucose itself is the "food" that is burned with oxygen in respiration, while CO2 and water are used by plants to make the glucose food by photosynthesis with the sun's energy.  Water and air could never be described as "food".  Food is what you get energy from by oxidation (burning it with oxygen).

The Cambridge dictionary disagrees with you.

I'm just saying it is more accurate to say that plants make their own food by photosynthesis, and that food is glucose, not "air and water", because there is no energy to extract from air and water: you have to put energy in to get the glucose (food), from which you can then get energy out.

There's a quote from Richard Feynman floating around in which he says that trees "eat air" to get sustenance. Over 95% of the trees mass is obtained from atmospheric Co2 and water.

One could also say that houses "eat bricks" to get sustenance, but that doesn't make it food.  Food is a source of both energy and building blocks.  Glucose provides both, but air and water provide no energy.

PS. If you don't believe me, check how many calories of energy are advertised on a bottle of water (or air for that matter). ;-)


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B-cell antibody production
I've just learned about B cells in immunology lectures and some things are not clear to me.
Here's what I know:
1) Apparently, each B cell produces a specific antibody, determined randomly at the birth of that cell.
2) A B-cell expresses 250000 different antibodies on its surface
My question:
Are both these facts true? If so, how can they both be true if each B cell only produces a single specific antibody?

The specificity of antibodies is answered below. However, I think the answerer misunderstood the "250000" number. What this actually refers to is the number of copies of the cell's antibody that are on the cell surface - each antibody-producing B cell has approximately 250000 antibody molecules on its surface. Each one of those copies is identical, all recognizing exactly the same epitope.

Statement 1 is true and Statement 2, false.
B cells, in the absence of antigenic stimulation express surface receptors(B-cell receptors or BCRs), which look like normal secreted antibodies (but these are membrane bound and not secreted). What is amazing is that each B cell at a particular time expresses the same B cell receptor - same as in, all receptors produced off one B cell are of the same type, say, antigen A. Now, another B cell produces another type of receptors, say, those which detect Antigen B. There is not just one BCR on the B-cell, but many - just like any other cellular receptor.
Now, there are many B cells in the body. Very much like antigen A and antigen B, there so many more antigens that our body can produce antibodies against. (The source I quote below says $\ 10^{12} $) It is a wrong notion that just one little B cell is doing that all. There are specific B cells for each specific antibody.
Each B cell isn't necessarily producing a unique antibody, but yeah, specific. It isn't unique since there must be a few more similar B cells producing the same receptor against antigen A. So it is not just one single B cell producing antibody against 'A'.
How does the body produce so many types of specific proteins?
Being proteins, they must be encoded by genes. B-cells while maturing, undergo a recombination (shuffling) of their immunoglobulin genes which leads to this differential expression of receptors, hence the diversity.
For further reading on genetics of immunoglobulins and the generation of diversity, I'd suggest - http://www.ncbi.nlm.nih.gov/books/NBK26860/

Also important to note is that there is significant antibody cross reaction with related antigens which makes the defense even more formidable!

Please see my comment above. Statement 2 is true, you just misunderstood the question.

@MattDmo the questions says, "250000 different". As you said, they arent different, but recognize the same epitope. The number meant something different, ill edit it in the answer! So statement 2 still remains false anyway for the 'different' part.

I'm not sure how "different" got in there, I suspect it may be a translation issue or something similar. I suppose the BCRs are "different" in that they're different individual molecules, perhaps what the OP meant to say. I don't know... :)

@MattDMo true that! :D Maybe that's what different meant!! I have edited the answer, but anyway could you pls paste a reference regarding the 250000. Like, the number of receptors expressed on the cell.

Statement 1 is true. Statement 2 is false.
My explanation is from the perspective of immune cell development:
The antibody-producing B cells are called plasma B cells. Plasma B cells are differentiated from a single naïve B cell (undifferentiated B cell). The naïve B cell have all the cell structure. However, naïve B cell can also be differentiated into memory B cells. When one single B cells receives differentiation signals from a T helper cell and differentiate into a plasma B cell, it grows bigger and starts proliferation. Thus you end up with many plasma B cells all coming from the same very first plasma B cell differentiated of this type. So one type of plasma B cells MUST present the SAME type of receptors on their surfaces.

Primary Reference: Cellular and Molecular Immunology, 8th ED.
Statement 1 is only part true. B cells produce b cell receptors of a specific paratope, that are randomly determined during maturation. The process of V(d)j recombination, where productive rearrangements in the heavy chain and light chain genes, produces a primary CHtranscript consisting of V-D-J-Cµ-Cδ. Enzymes in the matured B cell can differentially process the H-chain transcript into VDJ-Cµ or VDJ-Cδ which confer IgM or IgD-isotype BCRs. Mind you, during recombination and rearrangement, the extra DNA is deleted that would otherwise lead to other receptor possibilities, and signals are activated that prevent any potential recombination from taking place on another chromosome. This maintains a single antigen specificity on all clones of a given B cell!
The only difference between a BCR and an antibody produced by that B cell is the presence of a C-terminal membrane anchor region. How the protein is processed is determined by what proteins are processing the primary mRNA.
B cells retain in their DNA C regions for other heavy chains, however. In the presence of certain costimulatory signals, they can class switch to a different isotype that retains the paratope of the B cell. I explain this at length here.
I want to stress here that only the paratope or antigen-binding site is maintained, and B cells can produce a number of different isotypes of BCR or secreted antibody (though class switching is generally irreversible).
Statement 2 follows that due to the above statement, a B cell cannot express 250,000 different BCRs. Rather, they express one paratope across different isotypes. While I don't have the data in front of me, 250,000 may very well refer to surface density of BCRs on a given B cell.


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Is "A" inhibiting "B" and vice-versa, a positive feedback loop?
If product A inhibits the expression of product B, and product B inhibits the expression of product A, is this a positive feedback loop?
My thinking was to consider the scenario that A starts off high, and so B is naturally low. Then, a large amount of B is added to the system. This inhibits the production of A, so the level of A falls, and that in turn enables the production of more B, which further decreases the production of A and in turn further increases the production of B, in a never-ending cycle. This runaway amplification is characteristic of positive feedback loops, whereas negative feedback loops cause a return to equilibrium.
Yet the Wikipedia page says that, for it to be a positive feedback loop, it must be set up so an increase in A causes an increase in B.
So what is it?

This is probably not a biology question. The phenomenon you describe is a positive feedback IMHO.

In essence yes, because the more A, the less B, the more A.  In practice it would be a pretty unstable system so you'd end up with either all A, or all B.

The situation that you presented in which an entity A inhibits the production of another entity B which in turn inhibits A, is a positive feedback.
In a network path or a loop the overall sign of the loop/path is the product of the signs of individual edges (interactions). In this case it is negative times negative which gives a positive sign to the loop.
In simple words, a positive feedback should lead to an entity promoting its own production, directly or indirectly; it can auto-activate itself, activate an activator or inhibit an inhibitor. In this case A inhibits its inhibitor. Sometimes you can have large loops like this:

This is still a positive feedback.
Moreover, your case would exhibit (with the right parameters) the dynamic properties of positive feedbacks (such as bistability and hysteresis).
Sometimes people also refer to this type of positive feedback as double-negative feedback (which I personally do not like because it is misleading).

Positive and negative feedback loops may consist of a single component
  that activates and represses directly its own activity, respectively
  (Fig. 1B,C); or they may include several components and involve
  indirect interactions (Fig. 1D–G). The overall sign of a complex
  feedback loop (i.e. positive or negative) depends on the constituting
  elementary interactions (Fig. 1D–G).(1) For example, two mutually
  repressing components form a positive feedback loop (PFL, also termed
  “double-negative feedback loop”).

Mitrophanov & Groisman, 2008

However, not all positive feedbacks lead to runaway amplification. That is a property of an unstable system. The degradation of the molecules (or death) ensures a finite (and stable) steady state. Sometimes (with certain parameter combinations, as previously mentioned) positive feedbacks can have two stable steady states. Depending on the initial condition, the system can settle to either of the states. In your example the two states can be — High-A Low-B and Low-A High-B.

Further reading:
Mitrophanov, Alexander Y., and Eduardo A. Groisman. "Positive feedback in cellular control systems." Bioessays 30.6 (2008): 542-555.

Although the situation you describe involves amplification rather than homeostasis it is not a positive feedback loop under the standard definition of the latter:
In a a positive feedback loop:

An increase in A leads (directly or indirectly) to an increase in B.
The increase in B leads (directly or indirectly) to an increase in A.
This repeats step 1., leading to a further increase in B.

The second step is feedback, as it is affecting the initial propagator of the action, and positive, as it causes an increase in that propagator, A as well as target B.
Your model differs from this in two ways. First, only B is amplified — A is suppressed. Second, when A falls to zero B will reach a maximum, whereas in a positive feedback loop things theoretically keep on running out of control. (Yes they generally stop, but that is because of the intervention of something outside the simple A/B system.)
Some biological examples illustrate this difference (please, I am not concerned whether they are true or not).
An example of a positive feedback loop might be atmospheric warming (A) causing production of more water vapour (B) which has a greenhouse effect and increases atmospheric temperature (A), which...
An example of your model might be the territorial encroachment of smart and aggressive Homo sapiens (A) causes a decline in the population of H. neanderthalensis (B) which is then less able to resist Homo sapiens, leading to a further decline and eventual extinction of H. neanderthalensis.
At the end of the day, we are arguing about semantics. Your point of view would seem to be that the current definition of positive feedback should be extended to include your model. My point of view is that scientific terminology allows us to distinguish between similar but different things. So, (not meant as a serious proposal) one might include both effects under a broader description as ‘augmentative cycles’, including both the standard ‘positive feedback loop’ and your model, which could be called a ‘positive–negative feedback loop’ or a ‘terminal-positive feedback loop’ or whatever takes your fancy.
Postscript
There is a paper in Current Opinions in Cell Biology which discusses the type of motif you describe under the subheading ‘Positive Feedback Loops’, although also refering to them in the text and in the title of a table as ‘double-negative feedback loops’. As @WYSWYG mentions, the review he originaly quoted only uses the latter term, and has a diagram of two-component regulatory motifs, which I reproduce below, distinguishing this from positive feedback. This bears out what I wrote — not that your point of view is necessarily wrong — but that this is a question of semantics, and scientists find it useful to use different terms to distinguish different but related things. In any case, the authors of the review have done the reader a service in defining their terminology.

I disagree.  If you consider that increasing the presence of A increases the absence of B which increases the presence of A, then it fits your definition of a positive feedback loop.  In other words, the absence of B corresponds to an increase in the signal of its absence.  And what really matters is that more A leads to more A.  So it acts like a positive feedback loop because it is unstable.  Whereas a negative feedback loop would reach a stable equilibrium where more A leads to less A and vice versa.

@Kelvin -- I don't think there's much room to disagree. In the context of Biology a Positive Feedback Loop has a distinct definition; an increase in A leads to an increase in B, whose product or byproduct leads to an increase in A in an (IIRC) exponential fashion until a limiting factor is involved.

I concur with Kelvin. A inhibiting B and B inhibiting A is a positive feedback. @MCM there is a room to disagree because this answer is not correct. In a positive feedback, an increase in A should directly or indirectly lead to increase in its own levels. It can auto-activate itself, activate an activator or inhibit an inhibitor.

I think it's a matter of perspective: If you're more focused on the detail and technical definitions you see only the individual steps; but if you're more focused on the big picture, you look at the system as a whole.  And since biology itself focuses on big picture systems and outcomes (via evolution), it's probably more useful to think in the same way than get bogged down in largely irrelevant technical details and definitions.

@Kelvin Apologies to you and the questioner that my original answer did not address the question adequately. Late at night, but that's not an excuse. I have not changed my opinion over yes or no, but I have argued and discussed it in a manner that pays more respect to the questioner's arguments.

@David Even if you do not look at the big picture, a positive feedback does not mean that all the individual interactions should be positive in nature. Basic graph theoretical concepts (that are also valid for systems biology).

@David, no problem.  I think it's good that you keep your answer posted as is, because it does illustrate that the answer is not so clear cut.  In any case it's a very interesting question.

@WYSWYG — I'll look at the paper you cite, but it will take a wee while. Ultimately it may be a question of different usage among different communities. If dpdt is correct, someone should edit the Wiki entry. Easily done, whether or not you're signed up (although latter is preferable).

@David The paper actually calls the structure as a double-negative feedback (a terminology that I do not like because it is misleading). You can introduce another node in this network and a positive edge and this can no longer be called "double negative" but it is effectively the same. Double negative can also be thought of as two parallel paths of a negative feedback (imagine a situation in which A activates B and C and the latter two inhibit A). However, as per graph/network theory this structure would be called as positive feedback. I changed the reference actually. See that one.


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What happens to the human brain when unconscious?
What part of the brain gets affected and does it harm the brain? Thank you I just needed some extra info for a video I'm making.

unconsciousness is a broad term. Consciousness itself is a debated term too. Could you add some framework around it? Are you talking of physiological sleep, drug-induced reduced consciousness, unconsciousness caused by head trauma or otherwise?

Unconsciousness induced by head trauma and drug induced reduced consciousness

Those represent different mechanisms of loss of consciousness. Might be better asked separately. I think this is a bit broad as is. Have you googled this? What did you find?

I didn't find anything worthwhile,just something about weed and alcohol harming the brain

Check out the ascending reticular activation system

I did but it doesn't really answer my question, could you explain your point in reference to my question...Thanks!

We don't know. Disruption of cortex, thalamus, or brainstem structures (someone mentioned ascending reticular activation system) all can cause loss of consciousness. Different drugs have different mechanisms of action and some might cause different "types" of loss of consciousness - for example, ketamine and propofol seem very different, and trauma is a completely different issue.
The reason this is such a hard problem is that these structures are all interconnected, and there are likely proximate and distal causes of loss of consciousness. For an analogy, imagine you starve to death. You could make a convincing case that lack of food did it - so conclude food keeps you alive. But you weren't declared dead when you stopped eating; you were probably declared dead when your heart stopped. So conclude your heart keeps you alive. But in the middle somewhere, you had changes in ion concentrations in your blood, loss of glucose. So salts and sugars keep you alive. I can prove these are all true - take out your heart, you die. Stop feeding you, you die. Fill your arteries with KCl, you die.


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Effective population size of a metapopulation structured on a grid
Scenario
Consider a rectangular metapopulation consisting of $x \cdot y$ demes, where $x$ is the number of deme horizontally and $y$ is the number of demes vertically. The population size per deme is $N$ and the migration rate between any two adjacent demes is $m$.
Assumptions
I welcome any kind of assumptions for the purpose of this post.
Question
Can you help me to calculate an expectation of the effective population size $Ne$ of the whole metapopulation?

I am not an expert in this area but if the initial population in all the demes is homogeneous then you can consider the entire metapopulation as one unit with x × y × N individuals. If the sex-ratios are skewed then you may expect different dynamics.

Mating is not random in the total metapopulation, the $Ne > x \cdot y \cdot N$ even if (as I assumed in the question) all demes have the same size $N$.


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Why does pETDuet-1 only have one T7 terminator but two T7 promoters?
Why does pETDuet-1 only have one T7 terminator but two T7 promoters (vector map below)? Is there any advantage of having only one terminator? There are going to be two mRNAs transcribed from this vector: one polycistronic (with two genes) and one monocystronic. Also, presumably, there will be more RNA for the gene expressed from the second promoter compared to the gene expressed from the first. That could lead to higher expression of the second protein, provided that the RBSs are the same. Is there experimental evidence confirming/disproving that hypothesis?
Is there any advantage over using polycistronic mRNA vs moncystronic? Have Duet vectors been used for heterologous expression of long (>40kb) biosynthetic pathways?

Sounds like the manufacturer could answer these questions much better than we can.

Not sure it can. But waiting from reply from them as well.

One of the duet user manuals (https://www.emdmillipore.com/CA/en/product/pETDuet%E2%84%A2-1-DNA---Novagen,EMD_BIO-71146#documentation) says that they are useful for studying protein complexes. I'm assuming it allows for more optimal co-translational assembly processes, and possibly favours 1:1 stoichiometry.

@Jory But in that case why is there a second promoter?

OK so I didn't really read up on this last time I answered. As you said the downstream ORF isn't translated as much, so it wouldn't favour 1:1 stoichiometry. I have a feeling the expression of the shorter transcript is to compensate for this effect, but I'm not sure.

Can you add the plasmid map? Overlapping genes would actually cause lower expression.

@WYSIWYG Added vector map. The genes are not overlapping.

Apparently the expression of the second protein in a polycistronic mRNA is lower compared to a two-promoter system (Ref). Anyway, this still doesn't answer why a single terminator is present.

Could they be thinking that the transcription complex from ORF1 would fall apart when hitting an new transcription complex that is being assembled on promoter/RBS 2?

I think the presence of a single terminator is related to the high processivity of T7 RNA polymerase.
T7 RNA polymerase will transcribe a circular plasmid repeatedly before dissociating. The T7 terminator after ORF2 ensures that the transcript terminates efficiently.
Based on your schematic, these are the potential mRNA transcripts from that construct:
Transcript from T7 promoter #1
 5'---RBS---ORF1---STOP----RBS---ORF2---STOP---[T7 terminator]---3'

Transcript from T7 promoter #2
 5'---RBS---ORF2---STOP---[T7 terminator]---3'

It is doubtful that this vector will produce precisely equimolar concentrations of each protein product, however, the topology ensures that there is at least some of each protein produced. Of course, this could be tested empirically.
I hope that helps!


text:
What is the highest number of genes expressed under one T7 promoter?
What is the highest number of genes expressed under one T7 promoter? Also, what is the longest polycistronic mRNA? I am interested in the context of heterologously expressing biosynthetic gene clusters in E. coli.

does T2A/P2A peptides work for your case?

@aaaaaa Not really, since I would like to avoid adding extra sequence to the coding region.


text:
Why use RRBS (reduced representation bisulfite sequencing) instead of general BiSeq?
My understanding is that RRBS is used for certain cases. For example, WGBS (whole genome bisulfite sequencing) is limited by repeated sequences.
However, many papers seem to use both techniques.
What are the reasons researchers would use RRBS instead of WGBS?

The use of RRBS instead of WGBS is mostly about coverage vs. cost.
To confidently call methylation status at a cytosine residue, you need to have at least 10x sequencing coverage at that particular residue (ENCODE standards require at least 10x coverage for RRBS and 30x coverage for WGBS). To perform whole-genome bisulfite sequencing (WGBS) on large genomes (e.x. mammalian genomes) is very expensive if you are trying to achieve the 10x coverage requirement.
The principle behind RRBS is to improve the sequencing depth of cytosine-rich areas by enriching your sequencing library with CpG-dense regions of the genome. This is accomplished through the use of CpG-specific restriction enzymes like MspI, which recognizes 5'-CCGG-3'.
In RRBS, the entire genome is cleaved using MspI, leaving at least two cytosines per cleaved DNA fragment (one cytosine at either end of the MspI-MspI fragment). Sequencing libraries are constructed from size-selected MspI DNA fragments, bisulfite conversion is performed, and the converted library is sequenced. Specialized software is needed for alignment and analysis of the converted genome (e.x. Bismark, methylKit).
One of the major limitations of RRBS is that you will sequence only CpG-rich regions of the genome. Also, since the library is size-selected after MspI digest but before adapter ligation, you will never sequence any MspI fragments outside of your size-selection range (~40-250 bp).
I have analyzed both types of data and found that the RRBS data was very unsatisfying and didn't answer our research question. WGBS, though extremely costly, is a better option for a comprehensive cytosine modification profiling. There are publicly available RRBS and WGBS datasets at https://www.encodeproject.org/.
I must mention a critical limitation of both techniques in that neither RRBS nor WGBS can distinguish between the type of modification (methylation, hydroxymethylation, carboxylation, etc) present at a particular cytosine residue. RRBS and WGBS, which rely upon the ability of certain modifications to protect cytosines from deamination, can only be used to determine whether cytosine residues are modified or not.
Further reading:
I would suggest following this link for an exceptional technical and practical article in which the authors compared RRBS to WGBS: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4026711/
A more technical guide to RRBS: http://www.bioinformatics.babraham.ac.uk/projects/bismark/RRBS_Guide.pdf

can you add some references for others to understand your answer better? it would improve your answer - thanks


text:
How is bilateral asymmetry determined in embryonic development?
What are the mechanisms that ensure consistent left-right asymmetric placement of various internal organs (heart, liver, etc) or consistent left-right asymmetric development of paired organs (brain, etc.)? At what level, and how comprehensive is our current understanding about the underlying mechanisms?
(It seems to me a mirror image is just as good as the original one in terms of fitness and functionality. But why only one specific LR asymmetry prevails? Maybe this is a misguided way to think about the problem but that motivated to ask anyway.)

There are two morphogens called lefty and its antagonist nodal. Their signalling and interaction gives rise to the right-left asymmetry. However, to answer your question fully, one would have to look up on how exactly are lefty and nodal regulated.


text:
A state model of sodium channels
I am studying by myself Human Physiology.
I have encountered the following question:
In the following given model of sodium channel with 3 states open closed blocked (which I assume means inactivated), the rates of going from state to stated are given in the picture below.
a. Write matrix $Q$ for the system and find it's eigenvalues.
b. What does these values represent?
If anyone has an idea what is the required matrix I will be grateful.
Thanks!

Interesting question, looks like homework but i won't give you a minus. Can you describe full description, i mean, what  these numbers mean. Suppose it is charge need to be possessed to change the state, if so, i can do that, it looks like simple matrix model if we add some probabilities of generating charge. I also suppose that you need to get eigenvalues just to find stable system state and that's good.

I think Q is supposed to be an adjacency matrix that represents the graph in your diagram. Take a look here: https://en.wikipedia.org/wiki/Adjacency_matrix and here: http://www.cs.elte.hu/~lovasz/eigenvals-x.pdf

This is a homework question. You should at least make some attempt at the answer.

Thank you all. It is from HW I found in the net. So, no other information valid for me. I usually, of course, make a lot of self effort. But, sometimes, like in this case, even though you know you have the right tools, you just don't know where to start, especially, if not all the definitions are given... So, Thanks for your help and comments!

There is no standard notation called Q (matrix).
However in this case I think the matrix that they are referring to is the state transition matrix (similar to the adjacency matrix as mentioned by Justas in the comments, but with rates instead of just the connections). Basically you have three states (lets call them A, B and C) and there is a rate for transition from one state to the other. You can also represent these rates in the form of transition probabilities.
You just represent this as a 3×3 matrix. Put zero where there are no such transitions (for example there is no transition from A to A or A to C):
$$\begin{array}{|l|c|c|c|}
\hline
 & A & B & C \\\hline
A & 0 & 10 & 0 \\\hline
B & 100 & 0 & 50 \\\hline
C & 5 & 0 & 0 \\\hline
\end{array}
$$
You can then calculate the eigenvalue for this matrix. How to calculate eigenvalues and what is their significance is off-topic in this site. You can easily find out how to calculate eigenvalues. Their significance is something that is not that easy to understand but you can read more about that and can perhaps ask a precise question in Mathematics Stack Exchange. Basically, they tell you how the system proceeds in different directions (denoted by eigenvectors).

About the eigen-values, I know that if I view this matrix as a correlation matrix, then the eigenvalues are the variance values in the 3 element basis of eigen-vectors. Can I apply this character here? Anyway, Thank you!!

@user135172 just that this is not the correlation matrix. However, the concept of eigenvalues and eigenvectors is similar. Just like any linear algebra problem. If a matrix is a function applied to a vector (which may denote state of the system), then the eigenvalues somewhat denote the amplification factor of the matrix (along different directions).


text:
Do transcripts always start and end with exons?
I realized that in all cases of "RefSeq Genes" annotations of hg19 I looked at spliced transcripts start (and end) with an exon. From the annotation there is no evidence of any sequence upstream or downstream of these exons that remain in the nascent RNA.
Does this reflect the biology or is it just a consequence of the spliced read mapping? In other words: Do transcribed regions upstream of the first exon and downstream of the last exon exist in nascent RNA?
These regions, if they exist, cannot be called introns when those are defines as 'spliced sequences between exons'. How would one call them?

Welcome to BiologySE... I agree with what you say at the end (i.e. "I guess if you have an answer for 1 and 2..." - I would narrow your question to get an answer (it's easier to answer a simple, straightforward question instead of a broad question with 5 subquestions). You can always ask more questions - the more specific the better

Very Good (and well-researched) question!! I am looking forward to answers here

@TalhaIrfan Thank you. I condensed the text to the basic questions.

Could you explain what you mean by "spliced read mapping" please.

@David, spliced read-mapping refers to the mapping of spliced sequences (e.g. RNA-derived sequence reads) to an un-spliced reference (DNA of reference genome). The algorithm that performs this kind of task has to be able to accept long insertions on the reference sequence without penalizing those as mismatches in the alignment. These read-to-reference insertions (or reference-to-read deletions) are introns.

@Herr Jemine - I see. Thanks for the explanation.

Most (almost all, AFAIK) mRNAs and lncRNAs start with exons for the reasons already mentioned by David. In a typical splicing event, the nucleotide that is 5' to the splice donor site (lets call it pre-donor) and the one that is 3' to the acceptor site (lets call it post acceptor) are joined together and the intronic sequence between them is removed.
If you look carefully at the mechanism, you'll notice that the pre-donor (i.e. the 3' end of the first exon) renders a nucleophilic attack on the phosphorous of the post acceptor (i.e. the 5' end of the second exon) which leads to the release of the intron.

 Image courtesy: http://mips.helmholtz-muenchen.de/proj/yeast/reviews/intron/spliceo_splicing.html

Therefore there is always an exon preceding an intron, in the transcript.
However, RNA processing can occur via a different mechanism in which the ends are just chopped off. This happens in tRNA processing, where RNAseP chops off a portion 5' to the mature tRNA region, called the leader sequence (see the figure below). I am not aware of such a mechanism happening in the case of mRNAs but a possibility certainly exists. One possible reason for mRNAs not having this kind of mechanism is because their 5' end is capped (co-transcriptionally) and cleavage of the 5' would lead to loss of cap and destabilization of the mRNA (all guesses, though). Similarly, the 3' of mRNAs is polyadenylated which also renders stability to them. Many lncRNAs are also capped and polyadenylated but there are exceptions (mRNAs too).
Since this mechanism is not very common, there is no systematic name for the excised ends. For the case of tRNA, it is simply called the "leader" sequence.

 Image courtesy: Leigh & Lang, 2004

Thanks for your detailed explanation. As David and you point out it is very unlikely that the splicing mechanism would be able to remove sequences upstream and downstream of the first and last exon respectively. Another important hint is that 5' CAP processing occurs co-transcriptionally. This, and the fact that a method called 'CAGE' is used to identify promoter regions by CAP-specific sequencing of mRNA 5' ends gives some confidence that 5' removal of a 'pre-exon sequence' is likely not happening in 5' capped mRNA. Yet we have not discussed the 3' end.

@HerrJemine I don't really know a reason for why that is not common. Even for the 5' part this was just a wild guess. Do not take it as a fact. BTW, the 3' end has poly-A which is also critical for RNA stability. However an RNA can be stable in the absence of cap and poly-A tail too. Since such cases are also very few, we can guess that this could be a factor for low occurrence of end chopping in RNAs.

I found some hint on post-transcriptional 3' end truncation in alternative polyadenylation on wikipedia (second paragraph of intro and section 'Alternative polyadenylation'). I'd suggest that you add this aspect to your answer and I'll accept it.

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

As far as I am aware, transcripts always start and end with exons. The reasons I wouldn’t expect otherwise (apart from my observations when examining Drosophila transcripts) are given below.
As you will be aware, the spliceosome (at least for mRNA) is a highly sophisticated multi-component ribonuclear protein complex, and has functions to both splice out the intron and ligate the ends of the exon. The diagram below, from a recent review, suggests that recognition of the splice sites also involves the exon.

And the authors write (my italics):

“Base pairing between the 50 end of U1 snRNA and the first six
  nucleotides of the intron, and up to three nucleotides of the exon,
  provides the main driving force for specific sequence recognition.”

So I think that the contemporary machinery for removing mRNA introns requires exon mRNA to ‘get hold of’.
If such removal of introns could serve a functional purpose you would imagine the machinery would have evolved to accomodate such situations, but if the main function of introns is to allow differential splicing of exons that specify regions of the protein, introns at the end of a mRNA would have no purpose.
Another point to consider is the RNA-polymerase binding site on the DNA (the extended and complex promotor). If introns arose originally as mobile elements (something suggested by the authors of the review) then an insertion at the 5' end would likely have inactivated the promotor, so no transcript would have been produced.
Of course, Nature is full of exceptions, and it would be foolish to exclude that introns of the type you postulate do exist (in some galaxy far away...).

So why the mark-down? If you vote someone down have the guts and the decency to explain why. What is so reprehensible about the idea that splicing evolved because it allowed variants of proteins by differential splicing and the machinery evolved to deal with that? That although there are introns in the UTRs of a mRNA, which perhaps affect ribosome binding etc, these are likely to have been secondary. If you have a complex that needs to bind two parts of a rope before it can snip and join it, it makes perfect sense for it not to work at the end. Or don't you like my last para?

Didn't downvote, but I would guess that the theoretical and unsourced nature of this answer would play a significant role. Also, I find it strange why you would ask for someone to explain their downvote in such a defensive manner.

@MarchHo I won't shoot the messenger, but when you down-vote you are prompted to make a comment. It is most annoying to provide a structure-based answer to a theoretical  (more hypothetical) question and end up with -1 so a casual visitor would assume something basically wrong with your answer. My guess now is that the self-deprecatory and whimsical Star Wars reference was misunderstood as an attack on the question. Perhaps I'll change it to Shakespear when I add a pic of the spliceosome later to clarify my argument.

Your answer is touching the point but a little explanation and some diagrams would make it better.

@David Thanks for the answer to my question. I like that you pointed out the structural difference between a splicing between exons and a potential sequence removal upstream and downstream without a 'splicig partner'. By the way I didn't down-vote either, but my upvote did not count yet (missing exp.).

@WYSIWYG - I've added an appropriate diagram, and hardened up the argument. I didn't switch to Shakespeare, but the quotation I had in mind was "There are more things in heaven and earth...". For example this week's Nature reports that N6-methyladenine has been found in eukaryotes where it was thought not to exist. Never say never.

@David thanks. I have already given you a +1. I understand that there are many unknown mechanisms (and yeah I had seen that article too) but I think we can possibly justify why they are rare.

@David just as a point of clarification, users are under no obligation whatsoever to explain or "justify" their votes, up or down. The message you are referring to appears to low-rep or casual users of a site, but not to more experienced users, who theoretically understand the voting guidelines. I would also point out, as March Ho did, that lashing out in such a defensive and insulting manner, is neither useful to the overall tone of the site, nor likely to prompt anyone to explain or change their vote. Be nice.

@MattDMo OK, OK. However I did get such a message recently and I have been posting quite frequently. The message does imply that that is the correct etiquette, which longstanding users should have learned.  And btw, I do not consider my manner was insulting. It was forceful and direct, but no more. However when I explained a downvote re. another posting recently I got my head bitten off, so I guess one should just accept the guidelines are more honoured in their breaking.

Thank you for a great question.
I would like to start by clarifying some terminology.
First, nascent RNA refers to an RNA molecule that is currently being transcribed and has not been processed. Processing can include the splicing out of introns or polyadenylation at the 3' end, for example. Mature RNA is (typically) spliced and polyadenylated.
Second, the current consensus definition of an exon is a transcribed region of the genome whose sequence can be found in mature RNA species. Importantly, there are two classes of exons: non-coding exons, and protein-coding exons. I think you may have these confused and I have included an image to help clarify the distinction between introns, coding exons, and non-coding exons, using the human GAPDH gene as an example.

In the figure, the solid blue boxes represent exons, which are present in the mature mRNA. These exons can contain both protein-coding and non-coding sequences, as I mentioned. For example, exon 1 of GAPDH contains only non-coding information that belongs to the 5' untranslated region (UTR) while exon 2 contains both coding and non-coding sequence information.
When you view the RefSeq annotation, you are viewing all exons (coding-and non-coding). For some of the genes, such as GAPDH, the first exon will be non-coding. Other genes may begin with a coding exon (I have yet to find one and since the 5'UTR is involved in ribosome binding, I am doubtful that many examples exist).
The regions of an mRNA molecule beyond its coding sequence are referred to as the 5' and 3' untranslated regions (UTRs).
I hope this helps answer your question.

This does not actually answer the question. I think the OP knows about exons and UTRs. Their question is about why a transcript always has to start with an exon. Perhaps you can add some explanation for this point.

I have not heard of a classification such as coding and non-coding exons. You can have exons that have a part of the CDS and a part of the UTR.

Thanks for your reply. But my question did not concern whether exons contain coding or non-coding sequences. I am aware of the fact that exons can be part of the 5' or 3' UTR in the mature mRNA and also that there are non-coding RNAs in general. As @WYSIWYG pointed out I would like to know if the unprocessed, nascent RNA always starts and ends with an exon or if their might me a removal of 'head' and 'tail' RNA sequences that were transcribed from the DNA.

@WYSIWYG Thank you for your comment. Perhaps coding vs. non-coding is restricted to certain bioinformatics circles, which is where I came across the term. I think the 5'/3' UTR convention is more intuitive.

@HerrJemine Thank you for clarifying your specific needs. As someone pointed out above, only a very limited subset of RNAs are cleaved at the 5' end post-transcriptionally (I wasn't able to find any examples of mammalian RNAs that are cleaved at the 3' end). You may be interested in exploring a technique called NET-seq link. It demonstrates pervasive transcription outside of the transcriptional unit in yeasts, flies, and humans.


text:
What is a regulable promoter? And how does one regulate it?
I'm reading a patent where they (in S. cerevisae YNP5 strain):

downregulate the ERG9 gene by replacing the native ERG9 promoter with
  the regulable MET3 promoter

What's a regulable promoter and how does one regulate it? Must one add something to the broth to down-regulate the expression?
I don't see anything special in the culture medium.

I believe MET3 is repressed by methionine. The repression procedure (amount, concentration, delivery method, etc.) varies depending on the experiment.

Regulatable promoter means that the expression of the gene downstream of this promoter can be either induced or repressed. Met3 is an example of such regulatable promoters and it is regulated by the addition of methionine to the media. In yeast, Met3 gene encodes for ATP sulphurylase, an enzyme in the methionine biosynthetic pathway. Upon the addition of methionine, the gene driven by Met3 promoter is repressed, as its expression level is reversely correlated with the concentration of methionine added to the media (Figure from this paper):

It would be great if you can also briefly explain what the other "non-inducible" i.e. constitutive promoter is. That will clarify the difference.


text:
Does drinking alcohol having UTI may help to kill the bacteria?
It's well known that ethyl alcohol kills bacteria. And amount of alcohol in urine may be detected by EtG tests (as long as 3-4 days after intake).
Does drinking alcohol in reasonable amount by healthy individual (non alcoholic) having UTI helps to kill bacteria in the urinary tract (in order to get rid of them quicker)?

Nope, it does not. Alcohol is resorbed by the body and metablized. Additionally, to kill bacteria, you need 70% alcohol - which is a very bad idea to have this concentration in your body.

@Chris That looks like an answer disguised as a comment...

@RHA At the moment it's a comment. It doesn't have any references or whatever, maybe I find some time later to write a real answer. In this  case I will delete my comment.

Drinking alcohol does not seem to be effective treatment for urinary tract infections.
In this study, drinking 0.85 g ethanol/kg body weight (60 g ethanol or ~150 mL of 40% ABV spirit by a 70 kg person) resulted in 1 g of ethanol in the urine. 1 g of ethanol in 200 mL of urine = 0.5% alcohol.
According to CDC:

In the healthcare setting, “alcohol” refers to two water-soluble
  chemical compounds—ethyl alcohol and isopropyl alcohol...These
  alcohols are rapidly bactericidal rather than bacteriostatic against
  vegetative forms of bacteria; they also are tuberculocidal,
  fungicidal, and virucidal but do not destroy bacterial spores. Their
  cidal activity drops sharply when diluted below 50% concentration,
  and the optimum bactericidal concentration is 60%–90% solutions in
  water (volume/volume).

In this study, the minimal inhibitory concentrations (MIC) and antimicrobial effects of ethanol on Listeria monocytogenes in tryptic soy broth was determined. Ethanol at concentrations up to 1.25% did not inhibit growth, but growth was strongly inhibited in the presence of 5% ethanol.
According to BJPS, the minimal inhibitory concentration of ethanol is 4-9%.
A typical symptom of urinary tract infection is frequent and burning urination. Alcohol irritates the bladder, so it can make the symptoms worse (Mayo Clinic).

To begin answering your question,

Does drinking alcohol in reasonable amount by healthy individual (nonalcoholic) having UTI helps to kill bacteria in the urinary tract
  (in order to get rid of them quicker)?

We have to clarify that EtG detects ethyl glucuronide which is a byproduct of ethanol and remains in the urine for several day. The presence of ethyl alcohol in urine is significantly shorter somewhere between 8-12 hours (reference).
So what does this mean? Well, in order to "kill" the bacteria causing the UTI it must be exposed to a minimal inhibitory concentration of ethyl alcohol for a certain amount of time. Depending on the bacteria causing the infection this concentration and exposure time varies.
So if the concentration of ethyl alcohol was high enough and the flow of your urination and micturition long enough, maybe, but is unlikely to have much effect.

Good answer, but I would get rid of the last paragraph. The concentration of ethanol in urine is never going to be high enough to kill even a small percentage of bacteria present. In the United States, a person is legally drunk if their blood alcohol content (BAC) is between 0.08% and 0.1%, depending on the state. Even assuming that unmetabolized alcohol is concentrated 10-fold, and a person is very drunk with a BAC of 0.3%, 3% EtOH won't kill much of anything, especially if it's in a biofilm, as UTIs are.

@MattDMo i think this should be edited into the answer

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


text:
How do prions transmit their conformation to other proteins?
I was reading about prions and many sources say something to this effect:
"Prions may propagate by transmitting their misfolded protein state: When a prion enters a healthy organism, it induces existing, properly folded proteins to convert into the misfolded prion form. In this way, the prion acts as a template to guide the misfolding of more proteins into prion form." (Wikipedia)
How can/does a prion cause another protein to change its shape to match the prion's?

As the definition says, they act as a template - think "mold"

@Gerhard Yes but how exactly does a protein mold another? Does the prion bind to it forcing it to change its shape? If so, why does that happen? If not what does happen?

They don't misfold other proteins. They do that to other prionic protein molecules. An auto activation of sorts.

@WYSIWYG but if prions are misfolded proteins, that would mean that when they "misfold" other "prionic protein molecules", they're misfolding other proteins...in fact "The protein that prions are made of (PrP) is found throughout the body, even in healthy people and animals...The normal form of the protein is called $PrP^C$...[which is] a normal protein found on the membranes of cells."

Is there a confusion between "misfolded" and "denatured"? Although I'd agree that " misfolded" probably is not the best choice of word here.

@Gabriel PrPc is capable of switching but at a lower rate. This is accelerated in the presence of the alternative (beta-rich) state -- PrPsc (hence autocatalytic). I meant that the prion protein does not affect any random protein (which the statement in your question implies). The mechanism is mostly unknown. There are many other prion like proteins too, which are not pathogenic and such switches are involved in memory formation as well.

It is thought that infectious prions exist as clusters forming a crystalline structure. When a protein with the same primary structure is encountered but with a different tertiary structure, the normal protein undergoes a conformational change in order to integrate into the cluster. Presumably there are molecular forces involved that induce the conformational change.
http://www.rsc.org/chemistryworld/Issues/2005/October/prions.asp

Like other forms of amyloid, aggregated prions form a beta-sheet structure, where individual beta-strands are linked together by hydrogen bonds between their extended peoptide backbones, and hydrophobic interactions between the side chains on either side of the peptide backbone.  The beta-strand at each end of the beta-sheet has unpaired hydrogen bonds and hydrophobic side chains on the exposed end, which act as a template to link with an incoming prion.  Thus an incoming prion molecule can be unfolded as it joins with the exposed beta-strand, rather like a zip, and a new exposed edge is formed, waiting for the next prion to join.
This chain reaction is rather like normal crystalization, except that the process of aggregation also involves a conformational change from a folded globular molecule to an extended beta strand, which is catalyzed by the presence of existing beta-strands.  So the aggregation of prion effectively catalyzes more aggregation in a positive feedback loop: Once a seed has formed, the aggregation process will continue to unfold and recruit normal protein molecules.  Thus BSE/CJD was originally thought to be a virus as the aggregated conformation appears to be "infectious", passing its information about its aggregated state to a native folded prion molecule.
See for example:
http://www.annualreviews.org/doi/abs/10.1146/annurev.bioeng.4.092801.094202
http://www.annualreviews.org/doi/abs/10.1146/annurev.pathmechdis.3.121806.154326
http://www.nature.com/nrn/journal/v11/n3/abs/nrn2786.html
Or just google "prion aggregation mechanism review"


text:
What is the frequency distribution of each base in a DNA sequence?
Can we say that the frequency distribution of each base in a DNA sequence is equiprobable?

After the negative answer; i rephrase: Is there a use case in which the frequency distribution of the bases is equiprobable?

Just making an educated guess... DNA sequences are not random, thus the probability of any base being present at a particular location is not equal for all the bases.

My question was about their frequency.

? so should your question say "what is the probability distribution" or "what is the frequency distribution"? I guess I'm not following your question - can you help me out?

No. It is not. Very simple case - see GC percentage.

The frequency of the bases in the genome isn't equal to 0.25, the frequency depends on what kind of organism you mean. However let's take a look at some of them:

bacteria, most of the time we can see a bias towards some bases, this could be a GC bias, for example if the bacteria lives in extreme conditions, because GC can vorm three hydrogen bonds compared to AT which can only form two hydrogen bonds. So it's genome is more resistent to extreme temperature, less denaturation chance because the stronger bonds. In bacteria there are also some polymerases which have a bias towards bases, I won't discuss this in depth to keep it short, but from these two points we can already conclude the genome isn't random
Homo Sapien, for example we need CpG Island near our promotor region, to regulate transcription. Further we have higher GC% in gene coding regions to reduce mutation chance, because we don't want to mutate genes wich are needed for survival. Their are also some sequence which are very specific for example TATA box and Poly-A signal, if our genome would be random, so all base would have a frequency of 0.25 these specific sequence could occur at place where we don't want them or they wouldn't occur at places where we want them. We need a promotor (with TATA box) for RNA polymerase binding, if our genome was random we have a big chance we would have genes wihtout a TATA box so no transcription of these genes will occur

There are a lot  more properties of the genome we could discuss but I think this will give you enough insight why the bases in the genome don't have a equally alike frequence of 0.25
UPDATE:
in addition to your second question: I think the chance of finding a genome which has an equally alike distribution is very small. because almost all organisms have undergo evelution, and when we have evolution there is natural selection and natural selection relies on mutations. So (almost) all genomes have udergo mutation, however this sometimes can favor an equally distibution for example if our genome is AGAA and one A mutates to an G we will have an equally alike proportion. So to summarize: maybe you can take a look at NCBI and look for a gc 50% wich means 50% AT so 25% of each base. If you find a genome wich has exact 50% gc, you have found your answer

There are two main driving factors to consider here:
First, G/C and A/T represent complementary base pairs, as G is always paired with C (and vice versa), while A is always paired with T (and vice versa).  Therefore the frequency of G is always equal to C, and the frequency of A is always equal to T.
Second, G is paired with C via 3 hydrogen bonds, while A is paired with T via 2 hydrogen bonds.  Therefore, G/C bas pairs are more thermodynamically stable than A/T base pairs, and require higher temperatures (more energy) to be pulled apart during replication and also transcription.  For that reason, organisms that have a higher body temperature tend to have a higher proportion of G/C base pairs than those which have a lower body temperature.  For example, the G/C content is higher in thermophilic bacteria than in mammals, and higher in mammals than in plants.  This fine-tuning of the G/C content enables the optimum balance between stability and the extra energy required to pull the strands apart when required for replication and transcription.


text:
Taxonomic clasification of BLAST output
I am working with Next Generation Sequencing. I BLASTed the bovine de novo assembled contigs to the NCBI nucleotide database. I got a matching with a lot of species i.e. different kind of bacteria, viruses and mammals. I want them to group in a logical way i.e. all the bacteria in one group (bacteria group) similarly viruses and mammals. This is hard to arrange manually, can anybody help how I can arrange my vast number of species into some logical grouping. Thanking in anticipation.

I think this question is missing a few key details for it to be answerable. Do you have the NCBI IDs? A "logical way" will depend on protein context (membrane or globular, what is the enzymatic function, etc). Would the super-kingdoms be enough resolution?

This is basically metagenomics.
Congratulations, you already did the most time consuming step.
There are several ways to go from there, but I will talk about the one I know best.
There is the metagenomic analysis tool MEGAN. It can read your Blast Output, if it is in the correct format (normal XML or tabular) and will automatically do what you want.
For a start, just use the default values, for more help you an then go to the Community Website of MEGAN for help.
Analysing your data may take a little, depending on how many contigs you have. But if you were able to blast them in reasonable time it should be no problem. As I said, you are through the most time consuming step already.


text:
What did cyanobacteria gain from photosynthesis?
As far as I understand, cyanobacteria began performing photosynthesis long before plant cells as we know them arrived on the scene. But cyanobacteria do not seem to use polysaccharides in the same way as plant cells do (building materials, for example). So what evolutionary benefit did cyanobacteria gain from photosynthesis, that could have made them continue to produce oxygen generation after generation?

But cyanobacteria do not seem to use polysaccharides in the same way as plant cells do (building materials, for example)

The Calvin-Benson cycle produces glucose which is the starting
material for a lot of biosynthetic pathways including that of the
nucleotides (ribose from the pentose-phosphate pathway). Glycolytic intermediates are also involved in the synthesis of amino
acid, isoprenoids etc. Essentially all the carbon that cyanobacteria
contains comes from fixation of carbon (from CO2) via
photosynthesis.
Cyanobacteria also store starch as an energy reserve (Suzuki et
al., 2013). They also produce other polysaccharides (Phillipis
and Vincenzini, 1998). Moreover, the peptidoglycan cell wall has a "glycan" part which is a glucose derivative (a polysaccharide).
Moreover, cyanobacteria cannot produce ATP via the photosynthetic
electron transport chain in the dark and ATP in dark conditions
is generated (though significantly less) via substrate level phosphorylation in glycolysis.

Why do you think that the only benefit from photosynthesis is polysaccharide synthesis? Photosynthesis allows an organism to convert photons into chemical energy. That chemical energy can be stored as polysaccharides and used as a building material, but it can also just be converted into some other compound, or just used to run the organisms metabolism, which is the real significance of photosynthesis and the reason it supports most of the world's ecosystems.

I just know polysaccharides are a big deal. I am asking what other things cyanobacteria would use photosynthesis for that made it powerful enough to keep doing it at that scale.

Literally everything an organism does requires energy. Photosynthesis provides an electrochemical gradient that can be used to make ATP - which is then used in many, many chemical reactions throughout the cell as an energy source. DNA synthesis requires ATP. RNA synthesis requires ATP. Protein synthesis requires ATP.

So the ATP proces did exist in early cyanobacteria? Was it mitochindrial?

ATP is found in  all known organisms, so there's every reason to think it would have been. Bacteria don't have mitochondria, so no, it wouldn't have been mitochondrial.

Sorry, I meant that there was a photosynthetiv ATP, i.e. a more powerful one than in non-photosynthetic anaerobic organisms.

Mitochondrial oxidative phosphorylation and cyanobacterial photosynthesis are really two special cases of electron transport chains, which come in endless forms. They all follow the same principle, namely the transfer of high energy electrons redox acceptors, favourable reactions which release energy that can be harnessed. The donor of high energy electrons can be anything (NADH in oxphos or P700 for photosynthesis) and the electron acceptor just has to have a more negative redox potential than the donor. As long as the transfer of the electron from the donor to the acceptor releases..

energy, it can be harvested. In the case of photosynthesis, donor electron is only so high in energy because the reaction centers transfer energy to it from the sun's radiation. By contrast the donor electron in oxphos is high in energy initially, with the energy input coming from the initial synthesis of food molecules. There is really an amazing variety of these electron transport chains, which often use similar enzymes and electron transfer machinery, but have extremely varied high energy donors and oxidizing acceptors. It could be both are inorganic compounds like sulfur and nitrates.

Often the energy is used to pump ions across a membrane to synthesize ATP using the stored energy of the electrochemical gradient, as occurs in the mitochondrial membrane. In most bacteria this occurs in the cell membrane, which nicely illustrates the endosymbiotic (bacterial) origin of mitochondria. Some phototrophs transfer energy to electrons using the sun's radiation, but then simply use that energized electron to form a chemical bond (instead of pumping ions), which can be broken to release energy and the low energy electron. So, there even are ways to store food aside from sugar and ATP.


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CRISPR/Cas for editing the human genome
I know, that the CRISP/Cas approach for "cutting" the human genome is not completely suitable if we can't say not suitable at all. Because we have many repeats and this approach can bring to our genome additional breaks in DNA and after DNA-repair it causes unwanted mutations. If so, why scientist still continue to play with it (See the link, only 16% success chinese scientist got) ? And what other DNA-editing mechanisms we have? I might be wrong, so correct me please.
Thanks for explanation.
Chinese Scientists Defy Ethics, Double Down on Editing Human Embryos

It's not clear to me what you're asking. Editing the human genome is not at all the same as editing the genome of human embryos. The vast majority of all work with CRISPR is for editing genes in isolated human cells grown in laboratories, not in people. In the laboratory setting, 16% is an excellent success rate; before CRISPR, that number was much, much lower --- small fractions of a percent. Very few scientists "play with it" in human embryos, because of the major ethical problems involved. So are you asking about molecular biology or ethics?

Ofc about molecular biology.

In terms of molecular biology, how do you come up with the idea that CRISPR is "not suitable" ? The success rates and level of specificity achievable with CRISPR is pretty darn revolutionary. That's why scientists continue to play with it -- in the lab, and in model organisms. Using CRISPR in humans is a whole different question though, and most scientists would say it's too early for this yet; but this depends a lot on what specific application you're thinking of. For curing a recessive genetic disorder for example, 10% success rate might be fine. You need to be more specific I think.

I'm interesting in how we can avoid unwanted mutations that CRISPR/Cas causes.

Well that's an entire research field ... If you're asking about the current rate of off-target effects, that can be answered (Keith Joung's paper mention by Tom is a good example). But in the study you cite, I don't see any data on off-target effects. They only looked at the CCR5 locus, and 84% of embryos that were not "successful" were either not edited at that locus, or contained indels (unsuccessful homology repair). So that is more about efficiency of homology repair, which is unrelated to Cas9 specificity.

This question has been flagged for closing. I think if dshulgin and @roland bundle the comments into the question and an answer then this is a productive question.

Repeats aren't the problem with CRISPR. They stop you from editing repeats and repeat-like sequences, but in principle they can be worked around through careful design of guide RNA. CRISPR techniques right now do have problems with off target effects, but these problems may yet be solved. The CRISP/Cas approach to gene editing already has a lot of different variants and is under continual improvement, so there's no reason to think it won't get better. There's also no reason to think that the 16% figure there is only influenced by the use of CRISPR/Cas - it's an experiment with lots of other variables.

Could you be more clear? Under repeats i meant that our genome has thousands similar to our target sites and we have big probability of incorrect interaction with Cas protein. Can you explain your point in more details? Sorry that's still unclear for me.

"thousands similar to our target sites" is not true of all sites. It's true only of sites resembling repeats. Most sites in the genome can, for instance, have specific PCR primers designed against them. This would not be true if they all resembled repeats.

"can" but is that true in general? I mean, we should have big confidence of that to process CRISPR/Cas on human's genome.

It is true in general yes. But not in every case.

The accuracy of CRISPR to its target is not necessarily a problem at this time, because scientists have developed a more accurate version of it, CRISPR hf, which has almost no off target effects.  Here's a report on a study with a very accurate version of CRISPR


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Which mammals cannot synthesize taurine?
It is fairly common knowledge that domesticated cats cannot synthesize the compound taurine. Other mammals seem to be able to synthesize taurine from cysteine [source]. Are there other mammals that lack the ability to synthesize taurine?

Apparently cats can synthesize taurine from cysteine by the action of cysteine dioxygenase type 1 (CDO1) and glutamate decarboxylase 1 like (GADL1)/GAD2 enzymes. You can find out which organisms cannot synthesize taurine from cysteine from here. I don't know how to parse this data at this moment and I don't have the patience to manually check.

@WYSIWYG The Kegg map for taurine metabolism is here but if you compare felis catus with canis familiaris you find no differences in the enzymes supposedly present. If correct, this is consistent with Gwen's post.

@David Yes. Possibly the enzymes exist but are deficient.

It is a misconception that cats cannot synthesize taurine.
Cats can synthesize taurine, just like other mammals, but not enough of it to make up for an entirely taurine-deficient diet. Cats (and other mammalian carnivores) would have consumed a taurine-rich diet in the ancestral environment. It is only when they are fed vegetable/fruit/grain-derived foods that they show symptoms of taurine deficiency--because those foods have low quantities of taurine and other taurine precursors.
Other domesticated or captive carnivorous mammal species are at risk of taurine-deficiency if they are fed vegetable-derived foods, though members of the cat family (Felidae) are particularly susceptible.

Primary reference: This 2003 paper about taurine concentrations in animal feed, especially the 'Discussion' section.

Taurine  is  an  essential nutrient of cats because the rate of taurine synthesis from its dietary sulphur amino acid precursors, cysteine and methionine, is much less than the extent of loss through faecal bile
  acids  and  urine  (Knopf et al.,  1978).  From  an  evolutionary  standpoint,  taurine  was plentiful  in  the  diet  of  a  true  carnivore,  as  high  concentrations  of  taurine  are  found  in muscle  tissue. However,  as  most  domesticated  felines  normally  do  not  consume  living
  prey, they are at risk to become taurine-deficient if not adequately supplied in the diet.

This would be consistent with the Kegg entries for enzymes of taurine metabolism for dog and cat, as I mention in my comment on the question.

Perhaps you can give an example of a mammal that can actually not synthesize taurine i.e. it lacks the enzymes.

@WYSIWYG I am not aware of any.


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Abdominal muscles during inspiration/inhalation
During inspiration I am aware that the diaphragm and the external intercostal muscles contract, thus increasing the volume of the thoracic cavity.
However, I read on my textbook that during inspiration:

the diaphragm contracts, and at the same time the external intercostal
  muscles and one set of abdominal muscles help to raise the rib cage.

What I do not understand is the movement of the abdominal muscles, as I thought that they help in ventilation during exhalation (contract during exhalation, but not during inhalation). Is my assumption incorrect?

In inspiration just the external intercostal muscles contract and in expiration the abdominal muscles contracts, and the external intercostal muscles relax thus the diaphragm relaxes and curves upwards. Furthermore, also in expiration, the internal intercostal muscles contract

Also note that during Eupnea, in inspiration only the diaphragm and external intercostal muscles are "required"

@JordiZambrino Okay, so what is written on the textbook is incorrect right? And during expiration the diaphragm relaxes? Thanks.

Not necessarily, its just that my textbook states something different. And for your second point: The Diaphragm relaxes  and curves upwards, the abdominal muscles contract, thus pushing the Diaphragm upwards, consequently the External intercostal muscles relax and the ribs fall, in the end, the Internal intercostal muscles contract, hence pulling the ribs downwards


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Are there well studied examples of ERAD-mediated membrane insertion, especially from viruses?
Membrane insertion of transmembrane proteins typically requires highly hydrophobic alpha helixes at the N-terminus, N-terminal signal peptides, tail anchors, or a combination of the three.
Byun, H., Gou, Y., Zook, A., Lozano, M., & Dudley, J. (n.d). ERAD and how viruses exploit it. Frontiers In Microbiology, 5
These processes occur co-translationally and are mediated by the SEC translocon and associated factors (especially GPI anchor transferases for tail-anchored proteins). However, my undergraduate research is focused on a viral protein which appears to be translated in association with SRP and co-translationally translocated to the ER by the translocon, but is not anchored to the membrane during translocation. This was shown (by others) by purification of the ER-associated fraction of the protein and centrifugation showing the majority pelleted with the dense, soluble fraction. A small fraction pelleted with the low density membrane-associated fraction, and my research further suggests that this membrane-associated fraction is important for its escape from the ER.
The protein I am studying functions in the cytoplasm, not in the ER or in the secretory pathway, and therefore its escape from the ER is essential for its function. The typical ERAD pathway for soluble proteins involves retrotranslocation through the Sec61 translocon channel (the same channel involved in co-translational translocation, but with different associated factors during ERAD), but this would result in unfolding of the protein prior to its retrotranslocation to the cytosol. On the other hand, the ERAD pathway for membrane proteins (the "dislocation pathway") appears to dislocate folded membrane proteins while retaining most or all of their tertiary/quaternary structure (previous citation and below).
Avci, D., & Lemberg, M. K. (2015). Clipping or Extracting: Two Ways to Membrane Protein Degradation. Trends In Cell Biology, (10), 611. doi:10.1016/j.tcb.2015.07.003
Knowing that the protein is not co-translationally inserted into the membrane because only a small fraction is membrane associated, and suspecting that the small membrane-associated fraction is important for its escape from the ER because the dislocation pathway allows folded protein to exit the ER, my question is:
How could the retrotranslocation pathway for soluble proteins be disrupted such that it would (infrequently) result in membrane insertion of this protein at an early step in retrotranslocation-coupled unfolding, such that the folded protein could follow the dislocation pathway into the cytoplasm?
I would appreciate any examples of similar processes in viruses or in eukaryotes, but any speculation on possible mechanisms - based on understanding of ERAD but lacking supporting examples - would also be greatly appreciated.
As this is unpublished research, I can't be too specific about what virus or even what model system I'm working in, but if there is additional information that would help, I'll be happy to provide it - if I can.
One final piece of information that is also relevant is that I suspect the protein's transmembrane or membrane-anchored domain is very close to its C-terminus. Another important question that this raises is whether retrotranslocation exclusively proceeds by feeding the N-terminus of a protein into the Sec61 channel, or if the C-terminus can be fed directly into the channel? If only the N-terminus could be fed into the channel, this would suggest most of the protein would have to be unfolded before the C-terminal domain became associated with the channel and was able to become membrane-associated, which voids the main benefit of the dislocation pathway; maintaining the folded state of the protein.
If somebody can answer only this question regarding the translocon (and provide some relevant reading), I would be incredibly grateful because it would suggest further literature search to help support the mechanism I am proposing.
Thank you!

This question is very dense and difficult to answer due to it missing key information. I've left a longer comment as an answer, but this is likely to get removed. If you could address the issues raised then a more constructive answer could be given.

The observed events are as follows:

co-translational translocation into the ER (may or may not lead to membrane insertion). This is abolished by deleting SRP and translocon components in a cellular model, and the protein is destabilized in these gene deletion mutants.

Low amounts of steady-state protein (but some) is membrane-bound

The protein may become associated with (not dissociated from) the lumenal face of the ER membrane during a failed ERAD process.

The very C-terminal end of the protein probably doesn't have a transmembrane helix, the N-terminus has a 10-residue helix


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How does the introduction of clones would affect population genetics?
Since the ability to clone animals gives another way to preserve species, this could help for the breeding process. It has been noted that with the ability to clone animals, endangered species can have their population numbers increased in a short time. How would this impact these population from the perspective of genetics/ the gene pool?

Welcome to Biology.SE. Is this a homework question?

yes it is @Remi.b

The title should be the main question quickly phrased and should not set the field of study. Obviously saying "it is a biology question" is really not helpful as everything on this site is about biology.

Then, you should always show your effort to try to answer the question otherwise the question is off-topic. I will vote to close as you haven't shown your efforts to answer the question and will remove the vote close as soon as you added this info. You can have a look at the homerwork question policy here.

Since the ability to clone animals gives another way to persevere species, this could help for the breeding process. Cloning helps to be alternative route for the genetic line and helps with endangered species to be save and increased.

You should add this comment into your question so that others will directly see what efforts you've produced. Try to come up with as many thoughts as possible. Despite showing that you are not just asking for "someone else to do your job", it also helps to target what may be unclear to you.

@kristy I have prepared an answer for you but, as this is homework, I would like to see more effort from you, some tips for you to think about: How do you think cloning could be used? Why would cloning be useful? Where can cloning step in? Why would this impact genetic variation?

@Kristy The question is on the verge of being closed, if that happens I can't post the answer unless it gets reopened - that will only happen if you make some edits to show that you've given some thought to the reasons that cloning may or may not be useful. It's otherwise a good question and I'd like to provide the answer


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What's the exact meaning of and how to derive the coefficient of the path from sire to offspring?
According to Wright's Coefficients of Inbreeding and Relationship,
the coefficient of the path from sire to offspring is given as
$p_{o \cdot s}=\frac{1}{2}\frac{\sqrt{1+F_s}}{\sqrt{1+F_o}}$
But how to get this formula?
And what is the  exact　meaning of this path? It's the path from sire's genetype to offspring's corresponding genetype, or just from one allele in sire's locus to offspring's one allele in the corresponding?
Furthermore, Wright gives
$F_o=\frac{1}{2}r_{sd}\sqrt{1+F_s}\sqrt{1+F_d}$
From Wright's another paper Correlation and Causation we know
$d_{o \cdot s}+d_{o \cdot d}+d_{o \cdot \bar{sd}}=1$
in which
$d_{o \cdot s}=p_{o \cdot s}^2$,
$d_{o \cdot d}=p_{o \cdot d}^2$,
$d_{o \cdot \bar{sd}}=2p_{o \cdot s}p_{o \cdot d}r_{sd}$
Thus
$\frac{1}{4}(1+F_s)+\frac{1}{4}(1+F_d)+\frac{1}{2}r_{sd}\sqrt{1+F_s}\sqrt{1+F_d}=1+F_o$
$\implies F_s+F_d=2$
Since $F_s,F_d\le 1$,
we have $F_s=F_d=1$......
Incredible. But what's wrong with it?

I think it must be forwarded to Math.StackExchange rather than stagnant here

In The Method of Path Coefficients(Wright1934) Wright showed that the path linked two objects is the path linked their genetic constructions. Probabily "genetic construction" means sequences of loci rather than a locus. However, if we consider a genotype of a locus as a random varible, the total genetic construction of a germ cell will be a random vector. Hence something like correlation or covariance between genetic constructions of germ cells is usually in form of matrix or tensor rather than a real number $F$(which is Wright actually showed). Moreover, if germ cells are made of several chromosomes, the correlation between corresponded pairs of loci in two germ cells may be not equal to each other. (e.g. chromosomeI of $G_1$ and $G_2$ are from the same ancestor, but chromosomeII are not)

So I guess things were worked out in this way.
Suppose $G_1=(g_0,g_1,\dots,g_n)$,$G_2=(g'_0,g'_1,\dots,g'_n)$. If $g_i$ is mth chromosome's kth locus and original from an ancestor germ cell $G_A$. And let us only modify $G_A$'s mth chromosome's kth locus, then $g_i$ would get modified; $g'_i$ would also get modified if only $g'_i$ is also original from $G_A$; loci on $G_1$ or $G_2$ in other chromosomes or in mth chromosome but other loci would not get modified anyway.  Hence $\rho(g_i,g_j)=\rho(g_i,g'_j)$ for any $j \neq i$. Therefore $$cor(G_1,G_1)=cor(G_2,G_2)=\mathcal{I}\dots(1)$$ $$cor(G_1,G_2)=diag(\rho_{g_0,g'_0},\rho_{g_1,g'_1},\dots,\rho_{g_n,g'_n})\dots(2)$$
Since $1=\sum\limits_{i}{p^2_{G_1\cdot g_i}}+\sum\limits_{i\neq j}p_{G_1\cdot g_i}\rho_{g_i,g_j}p_{G_1\cdot g_j}$ and by $(1)$, $1=\sum\limits_{i}{p^2_{G_1\cdot g_i}}$. hence $p_{G_1\cdot g_i}=\frac{1}{\sqrt{n}}$ if we weighted every $g_i$ the same. And $p_{g_i\cdot G_1}=\rho_{g_i, G_1}=\rho_{G_1, g_i}=p_{G_1\cdot g_i}=\frac{1}{\sqrt{n}}$, Analogously to $G_2$.

Therefore $\rho_{G_2,G_1}=\sum\limits_{i,j}p_{g_i\cdot G_1}\rho_{g'_j, g_i}p_{G_2\cdot g'_j}=\sum\limits_{i}\frac{1}{n}\rho_{g'_i, g_i}=E(\rho_{g'_i, g_i})$. i.e. in this case the correlation between germ cells are just the mean of correlations between their corresponding loci.
So similarly the coefficient of the path from a sire to its offspring is also the coefficient of the path linked their genetic constructions. and in detail is from sire's  genetic construction to its components, then from components to offspring's construction's components, then to offspring's construction.
The derivation was given in the article. In fig6 the coefficient of the path from a sire to its offspring can be gotten from the product $p_{o \cdot G_s}p_{G_s \cdot s}$.
$p_{G_s \cdot s}=\sqrt{\frac{1+F_s}{2}}$; $p_{o \cdot G_s}=\sqrt{\frac{1}{2(1+F_o)}}$.
So $p_{o \cdot s}=\frac{1}{2}\frac{\sqrt{1+F_s}}{\sqrt{1+F_o}}$

If $F_s=F_d=F_o=r_{sd}=0$, then $d_{o \cdot s}=p^2_{o\cdot s}=d_{o\cdot d}=\frac{1}{4}$, $d_{o \cdot \bar{sd}}=0$ . Thus
$d_{o \cdot s}+d_{o \cdot d}+d_{o \cdot \bar{sd}}=\frac{1}{2}<1$. This result means  parents' genetic constructions are not sufficient to determine their offsprings'. But it is very normal the same parents have both sons and daughters. Which causes their difference in gender are random factors.