human biology - What causes fingerprints to form and why is the pattern formed unique?

I would say genetic diversity is the primary reason which results in other reasons that you are looking for. At the lowest level, random crossing over at prophase I, random separation of homologous chromosomes at anaphase I, random separation of sister chromatids at anaphase II, and random fertilization: one sperm fertilizes one egg randomly.

The skin is developed from ectoderm so need to look at the formation of embryonic disc and specifically to the genesis of germ layers: ectoderm.

However, I would stick to the primary reasons, since it is extremely difficult to visualize the given formation - actually we do not have resources for it at the moment.

---

Very good question the last part. I have an intuition that skin develops randomly because of the above reasons. You would also need a lot of memory to make identical skins for twins! It has not been useful to have identical fingerprints between two people so evolution has not resulted into it.

Feeling surfaces and gripping are movements - not much space taken things, in contrast to the memory needed in storing the exact surfaces of skin from one generation to another. - Learning is a way to save resources here and it is a lot more efficient and than storing static information to species from one generation to another.

molecular biology - Intrinsically disordered proteins as potential drug targets

IDPs are indeed attractive drug targets and there are ongoing efforts to develop drug molecules that block interactions between a disordered and a structured protein. According to this relatively recent paper, however, these efforts have not brought a drug on the market, yet.

A few promising studies have shown drug-like molecules that inhibit protein-protein interactions based on intrinsic disorder of one of the partners and target:

• oncogenic fusion protein EWS-FLI1 and RNA helicase A. A small molecule has been found that targets the disordered region of EWS-FLI1, blocks the interaction with the helicase and inhibits growth of Ewing's sarcoma.




• p53 tumor supressor and its interactor Mdm2. Mdm2, by binding to an intrinsically disordered region of p53, targets p53 for ubiquitination and also causes it to be transported out of the nucleus. Promising small molecules have been found that associate with Mdm2 and thereby block its interaction with p53.




• c-Myc oncoprotein and the interaction with its partner Max protein. This study demonstrates two small molecules that bind to c-Myc and stabilize its disordered conformation, which inhibits its interaction with the Max protein.

The challenge in targeting protein-protein interactions for therapies stems largely from the fact, that the protein-protein contact surfaces are much larger than those involved in protein–small-molecule interactions (1,500–3,000 Ų and (300–1,000 Ų, respectively) [2]. They are often flat and have no defined binding pocket. Also, IDPs often don't bind natural small ligands, that could act as starting points in developing drugs.

You may find this paper helpful:

Metallo SJ, Intrinsically disordered proteins are potential drug targets, Curr Opin Chem Biol. 2010 14(4): 481–488.

BTW: for a comprehensive, manually curated list of disordered proteins and regions, please check the Database of Protein Disorder.

molecular biology - The effect of the start codon GTG on translation in E. coli

The NCBI translation table translates all alternative start sites as methionines. To my understanding, all translation is initiated by the fMet-tRNA. I don't know if there are any exceptions to this rule.

Regarding translation efficiency, I only found a 1985 paper in PNAS (Reddy et al, PNAS 82:5656-60), in which they compared the translation efficiency of adenilate cyclase's own UUG start vs GUG or AUG, obtaining a translation ratio 1:2:6 UUG:GUG:AUG, suggesting that AUG is the most efficient one, followed by GUG. Also, Romero and Garcia, FEMS microbiology letters 84:325-330 (1991) compared the efficiency of AUG vs AUC, AUA and AUU, showing a much lower efficiency for those codons, but they did not compare it to GUG.

Why is DNA replication performed in the 5' to 3' direction?

DNA replications needs a source of energy to proceed, this energy is gained by cleaving the 5'-triphosphate of the nucleotide that is added to the existing DNA chain. Any alternative polymerase mechanism needs to account for the source of the energy required for adding a nucleotide.

The simplest way one can imagine to perform reverse 3'-5' polymerization would be to use nucleotide-3'-triphosphate instead of the nucleotide-5'-triphosphate every existing polymerase uses. This would allow for a practically identical mechanism as existing polymerases, just with different nucleotides as substrates. The problem with this model is that ribonucleotide-3'-triphosphates are less stable under acidic conditions due to the neighbouring 2'-OH (though this obviously only applies for RNA, not for DNA).

So any 3'-5' polymerase would likely need to use the same nucleotide-5'-triphosphates as the 5'-3' polymerase. This would mean that the triphosphate providing the energy for addition of a new nucleotide would be on the DNA strand that is extended, and not on the newly added nucleotide.

One disadvantage of this approach is that nucleotide triphosphates spontaneously hydrolyze under aqeuous conditions. This is no significant problem for the 5'-3' polymerase, as the triphosphate is on the new nucleotide and the polymerase just has to find a new nucleotide. For the 3'-5' polymerase spontaneous hydrolysis is a problem because the triphosphate is on the growing chain. If that one gets hydrolyzed, the whole polymerization needs to be either aborted or the triphosphate need to be readded by some mechanism.

You can take a look at the article "A Model for the Evolution of Nucleotide Polymerase Directionality" by Joshua Ballanco and, Marc L. Mansfield for more information about this. They created a model on early polymerase evolution, though they don't reach any final conclusion.

marine biology - What would be the best dredging/trawling tools to collect macrofaunal priapulids?

My first question would be is there any external indication of them such as air holes or anything else that you can assess from the surface? I would think that if there is, you could use transects and a quadrat. If you need to dive for them you can obviously make a weighted quadrat by filling pvc pipe with sand and gluing it together. Then take an area and lay out transects and and randomly choose different lengths along each transect to put the quadrat down and that will give you an estimate of density. If you actually need to extract them from the mud, I imagine that varies by species and you could make due with a grab sampler of the right size, something on the small end I would imagine. That way you aren't dredging up tons and tons of mud to sort through. If they are even rarer than I imagine, you could just lay out transects and swim them until you see something and do away with the quadrat. Hope this helps.

genetics - How to create a collection of anonymous sequences for teaching and testing?

Here is the approach I ended up using, in part thanks to all the contributions here.

The associated R script is below or can be downloaded from:

BOLDS SEQUENCE RECOVERY

This creates 999 unique sequence files in plain text, with each sequence being identified to species level and few species being found across more than one sequence.

It also creates the matching answer key.

You can start at a random location to so that files change every year/group.

I used R to query the BOLDS database (Barcode of Life), to download a file and to split this huge file into separate sequences.

Here is the R script

rm(list=ls())

complete<-"http://services.boldsystems.org/eFetch.php?record_type=full&id_type=sampleid&ids=(*)&return_type=text"
write(complete, file="your location on disk")

rm(list=ls())

sequences.id<-data.frame("file.name", "recordID", "genus_name", "species_name")
write.table(x=sequences.id, file="sequences_id.csv", append=F, sep = ",", row.names=F, col.names=F)



set.seed(10)
start<-sample(1:1000, size=1)

i<-start
k<-1

while(k < 1000){

  sequences<-read.delim(file=complete, skip=i, nrows=1, header=F)
  sequence.compare<-read.csv(file="sequences_id.csv", skip=k-1, nrows=1, header=F)

  if(! is.na(sequences$V24)){
    if(as.character(sequences$V24)!=as.character(sequence.compare$V4)){
      writeLines(text=as.character(sequences$V55), con=paste(k, ".txt", sep=""))
      sequences.id<-c(k, sequences[,c("V3","V22", "V24")])
      write.table(x=sequences.id, file="sequences_id.csv", append=T, sep = ",", row.names=F, col.names=F)
      print("kept")
      k<-k+1
    }
  }
  i<-i+1
  print(paste(k,"/", i))
}

neuroscience - Is the squid giant axon the fastest conducting unmyelinated axon known?

The conductance velocity in the unmyelinated axon has been calculated and measured to be proportional to the square root of the axon diameter (see for example: Rushton, 1951). Since the giant axon is, well, giant, it conducts much faster than others. AFAIK, all other large neurons studied are myelinated. Maybe try to find bigger squids! ;)

neuroscience - Is there a biophysical causation from local field potential (LFP) to spikes?

Many experiments showed that neurons tend to fire at some phase (usually trough) of local field potential (LFP) oscillations, such as theta or gamma rhythm. LFP is supposedly generated by a population of neurons with coherent currents induced by spikes. So there is a causal link from spiking activity to LFPs. Is there also a significant influence on the spikes directly from LFP (not via the hidden spikes that generated the LFP)?

Is it just an epiphenomenon, and or is there a possibility that it is partly a serious mechanism for neural computation?

gel electrophoresis - What would cause bands to appear lower on a nonreduced SDS-PAGE gel?

The major effect of changing the redox environment of a protein is the formation or breakage of disulfide bonds. Under sufficiently reducing conditions no disulfide bridges will be present, while under oxidizing conditions your protein will form disulfide bridges if it has the ability to do so.

Disulfide bridges can significantly change the tertiary structure of a protein and change the behaviour of the protein on a gel that way. Generally, disulfide bridges should make the protein more compact and make it run faster on a gel.

It is therefore plausible that your faster band is the protein with disulfide bridges, which will appear to run like a smaller protein as it is more compact. The slower band then is the protein without disulfide bridges which can adopt a more extended conformation.

I'm assuming you're talking about native gels here, as denaturing gels usually contain DTT or beta-mercaptoethanol in the sample buffer to reduce any disulfide bonds and avoid this effect altogether.

nutrition - What does the term 'bioavailability' mean?

Bioavailability is a concept which applies to nutrients and drugs which pass through first-pass metabolism, i.e. orally (and to some extent nasally) consumed substances. Anything absorbed in the gut first passes through the liver before reaching the rest of the circulation, and both the gut and liver may metabolise it to some extent. The liver in specific has the powerful Cytochrome P450 system, a huge variety of enzymes to break down all sorts of substances, although in some cases it can actually produce more active or even toxic forms instead of breaking them down.

This can lead to drastic reductions in the amount available in the systemic circulation after oral administration. E.g. propanolol (a beta blocker) needs to be given in 100mg doses orally while intravenously (avoiding first-pass metabolism) only 5mg are needed.

physiology - What is the effect of exendin on beta-cells?

I recently tested exendin on INS1e cells in an Edu incorporation assay (similar to BrdU incorporation) to observe if this compound induces proliferation of the cells. Compound incubation was for 24 hours. I saw no incorporation of EdU with this compound over untreated levels. In addition, during the assay I look at overall cell number with a DNA stain, but I did not see any effect on total cell number with exendin (ie, if it were toxic the total cell number should go down which I have observed with other compounds).

nutrition - What are minerals (other than zinc) that the human body cannot store

Essential "minerals", i.e., metal cations are magnesium, zinc, iron, potassium, sodium, manganese, molybdenum, selenium, cobalt, copper and even calcium, as we lose a tiny amount of it through urine and sweat. They are all "stored" in some way, but only temporally, so some amount has to be taken up daily. It would show only weeks later, however, if you have a deficiency.

Wikipedia adds phosphorus and iodine but they are not metals.

human biology - When to measure resting heart rate and blood pressure for following day-to-day trend?

I would be very surprised if the time of day made a difference. I've personally never heard mention of such a phenomenon in discussions with intensive care practitioners (where of course HR and BP are measured constantly). However this is only the case during rest, this paper (on horses) suggests that there is some difference in HR and BP after exercise depending on time of day.

You are right to attempt to control it however, I suppose. It doesn't matter when you take your measurements, as long as you take them at the same time each day.

I would say that one of your first suggestions of taking the measurements almost immediately after waking. Of course you would have to be relaxed as you assembled your equipment then rest for a few minutes in the posture you have chosen (this will matter) before taking the measurements but otherwise I can't see you having too great a problem.

zoology - Do cows produce milk excessively?

The average domesticated dairy cow produces far more milk than would be required to feed their calf. All cows, wild and domesticated, will only lactate in the period between their calf's birth and weaning. Milk is calf-food, and when there's no calf, there's no evolutionary advantage in producing milk.

On dairy farms, cows are milked twice daily, from spring (when they give birth) until late autumn. This mechanical milking 'fools' the cow into continuing to lactate. When a cow has stopped lactating, they will only start again after giving birth. This means that you can't just start milking a cow and expect to get milk.

Generally farmers milk their cows from spring (birth) until late autumn. The reason for stopping in autumn is simply because the grass grows much slower in winter, so there isn't enough food to support lactating cows. However, it is entirely possible to milk longer than a year; I know of farmers who milk their cows continuously for two years. These farmers will have to purchase a lot of supplimental food (like hay or silage) during winter. The advantage of milking for longer than one season is that the cows do not have to give birth every spring, but instead only every second spring.

I believe (but can't guarantee) that in winter, most milk purchased in a shop comes from the opposite hemisphere. I do know that here in New Zealand, we export a lot of milk to northern-hemisphere countries.

If you were to suddenly stop milking a cow, they might get sick but generally they will survive. It's still something to avoid!

I do not have an 'official' source for these facts. However, I grew up on a dairy farm, so this was my life.

dna - What is the contribution of viruses to the evolution of mankind?

Human endogenous retroviruses (HERVs) have been an interesting (and expanding) topic of research in evolutionary biology and medicine. A retrovirus has an RNA genome in the virus particle, and integrates with the host cell's DNA upon infection to hijack the transcription/translation machinery and produce copies of itself. For this reason, it is not possible to "cure" a cell of a retroviral infection; instead all of the infected cells must be destroyed or commit suicide through apoptosis, also known as programmed cell death. Common examples of retroviruses include HIV (causative agent of AIDS), HTLV (human T-cell leukemia virus), and the Hepatitis B virus.

Viral infections are most common in somatic cells, where any change in the DNA sequence is not passed along to the next generation. However, any retroviral infection in germ cells (eggs, sperm, and the progenitor cells that make them) could be passed on to the next generation, and over time spread throughout a population. Some HERV fragments are apparently completely inactive (at least as far as we are able to tell currently), and are a possible source for some of the so-called "junk" or non-coding non-regulatory DNA that makes up a significant part of our genome - one estimate claims that 8% of our genetic material is of retroviral origin. Other HERVs have been implicated in various pathological conditions, including multiple sclerosis.

I'm not sure that any human genes have been found that were picked up by a virus, stayed with it for a time (and potentially mutated), then was reinserted back into the human genome, if that's what you're asking. There are certainly many cases of genes and gene fragments (coding regions, promotors, enhancers, etc.) being moved around as a result of viral integration/splicing. http://en.wikipedia.org/wiki/Endogenous_retrovirus#Genome_Evolution is a large, rather well-written section with numerous links to the primary literature showing how our genome has been altered over the years by HERVs.

Defining paper(s) in epigenetics - Biology

I understand that Robin Holliday was the first to discuss the possible role of DNA methylation
in the control of Gene expression. In his paper "The inheritance of epigenetic defects"
he presents what is one of the first modern formulations of what we now regard as epigenetics. The term "epigenetics" itself was coined by Conrad Waddington although this predated our modern understanding of heredity.

Holliday, R., The inheritance of epigenetic defects, 1987, Science, 238, 4824

gel electrophoresis - Alternatives to ethidium bromide for staining small nucleic acids?

While ethidium bromide works well for staining larger single-stranded RNA or double-stranded DNA molecules, it doesn't stain smaller nucleic acids very well. I observed that at around 20 bases and below single-stranded nucleic acids are difficult to see with EtBr-staining unless the nucleic acids are highly concentrated.

What are good alternatives for observing small nucleic acids in polyacrylamide gels? The main consideration would be ease-of-use, and it should be possible without any unusual equipment.

speculative - Why Didn't Evolution Cause the Human Body to become Streamlined?

If streamlining makes movement/locomotion quicker and easier, why didn't the apes evolve into life-forms that had streamlined bodies (much like fish)?

As with everything in Evolutionary Biology, you must ask yourself: Gain vs. Cost?

In your specific case, the Gain is very little. Air isn't nearly as dense as water, so a streamlined form won't show a major benefit unless the organism is traveling very, very quickly. This is why you see it in birds; raptors can travel over 100mph while diving, and at those speeds small changes in drag can mean the difference between dinner and starving. Smaller birds often make very quick turnabouts and changes in direction mid-flight where, again, small changes in efficiency can mean the difference between life and death. The cost was is worth it.

For apes and monkeys, moving very quickly isn't a case of living or dying. That's what we evolved opposable thumbs and prehensile feet(/tails) for. You don't need to run fast when you can climb a tree and simply get away from any predators on the ground. After we came down from the trees permanently, our larger brains allowed us to use tools to fend off predators - which, again, is much simpler than evolving an aerodynamic form that won't make a difference until you're running at the speed of a car.

So, in lieu of becoming a land-shark, we have hands that can use keyboards and minds that can invent the keyboard. Unfortunately, while the gains are many, the costs do include both a very long period of time where humans are helpless without parents, and an absolutely terrible form of locomotion with our upright stance on forward-pointing knees. Though you won't catch Cheetahs digging sewers anytime soon.

genomics - Sequencing the genomes of polyploid organisms

I've done some transcriptomics work in the past with a polyploid organism, and this presented some unique challenges in the data processing and analysis. Since then, I have been brainstorming about the technical challenges one may face when sequencing and assembling the genomes of a polyploid organism. As far as I am aware, there are no polyploids whose genomes have been sequenced.

If one wanted to sequence, for example, a tetraploid organism, one approach would be to prepare and sequence all of the DNA together and then rely on post-sequencing analysis to tease apart the two co-resident genomes. However, it would be difficult, if not impossible, with this approach to distinguish inter-genome variation from intra-genome variation.

An alternative approach would be to isolate DNA from both co-resident genomes separately, and then sequence and assemble the genomes separately, so that inter-genome variation and homology need not be considered. However, I'm thinking at a very high level and have little intuition as to the technical feasibility of this approach. When there are two or more co-resident genomes, is it be possible to isolate DNA from only one of those genomes? What would this rely on (for example, would thorough cytogenetic/cytogenomic characterization help)? If this task is not possible, what types of limitations must be overcome to enable it?

Is the "Great Pacific Garbage Patch" beneficial for marine wildlife?

There are very few things in the world that aren't beneficial to some lifeform. Even if you were to, say, spill a mixture of persistent broad-spectrum poisons on an area that killed off 99.9% of all species there, the remaining 0.1% that did survive would benefit from the lack of competition.

The "great garbage patch" is hardly so extreme a phenomenon, but similar effects can be seen there: some species suffer, others benefit. In that sense, it's no different from any other changing habitat.

From a conservation viewpoint, there are two issues here that one might find worrying:

1. First, novel, extreme or rapidly changing habitats tend to have lower than average biodiversity, at least initially: the few species that thrive in the new environment tend to proliferate at the expense of others, forming a relatively simple (and often not very stable) food web. Of course, over evolutionary timescales we'd expect the surviving species to adapt and diversify and the ecosystem to settle into a more stable state, but that tends to take a long time compared to the human timescale on which such novel habitats are created.




2. Also, the effects of the "garbage patch", or ecological changes in general, are not limited to the directly changed area. Creating a new habitat in one part of the ocean is one thing; disrupting the ecosystem of the entire ocean is something else, as doing so leaves no unaffected refuge for the species that are harmed.

   
   
   

   For example, fish, birds and marine mammals passing even just occasionally through the patch might end up swallowing plastic and accumulating it in their gut; meanwhile, even for less motile organisms, the patch might act as a population sink, depleting their population density in nearby areas. And if some organisms, such as the sea skaters mentioned in the article you cite, thrive in the patch, the increased population will likely spill into other areas, potentially harming their competitors or prey species.

As for why the news are focusing on the beneficial effects of the garbage to the sea skaters, well, that's what the study that prompted this current batch of news stories was about. It's also seen as newsworthy precisely because it seems so unexpected: we've all heard stories of plastic flotsam harming wildlife, so finding out that some species are actually benefiting from it has some "man bites dog" style news value.

Also, it's a lot easier and more convincing to observe something that is there than something that isn't: we can directly see these insects laying their eggs on the bits of plastic, giving direct and incontrovertible proof of a causal connection. Meanwhile, simply observing that some species used to be more common in the area of the patch than it is now is a lot less direct and informative: we might never be sure just how the patch is harming that species, and it always leaves skeptics an opening to claim that the decline might be caused by other factors. With the sea skates, on the other hand, we're not just observing the effect, but the direct mechanism as well.

homework - What is the main general difference between Mitosis and Meiosis?

I found such a clause:

The general principle is that mitosis creates somatic cells and
meiosis creates germ cells.

However, I cannot agree. Each gametogonium needs to go through mitosis before it can enter meiosis I. So in that case mitosis is happening with germ cells so the clause is false.

I would rephrase the sentence to be

The general principle is that meiosis creates only germ cells with the possibility of a decrease in chromosome number, while mitosis can create both somatic and germ cells while the ploidy stays constant.

Ok, not perfect.

How would you say the main general difference between mitosis and meiosis?

immunology - How does herpes (HSV) infection suppress HIV?

Alright, having read the citation linked, and doing a little poking of my own, here's my approach at an answer:

Some human herpes virus infections may compete with HIV infection. Essentially, some strains (not the ones you normally think of) infect CD4 cells - the same cells targeted by HIV. These strains down regulate transcription in CD4 cells, which in turn interferes with the HIV infection process. This pertains, it appears most notably, to HHV-7.

However the actual impact on HIV disease isn't clear. Strain competition triggers some fascinating evolutionary pressures, but HIV is notoriously prone to mutation, and competition for CD4 cells might not impact HIV infection on a clinical - rather than microbiological - scale.

Additionally, the two most commonly thought of forms of herpesvirus infection, HSV-1 and HSV-2 are associated with increased acquisition of HIV infection. The clearest reasons for this are genital lesions and inflammation at the site of HIV infection. There's also some interesting dynamics in play for active coinfection, such as the impact of acyclovir treatment for HSV impacting HIV, or HAART treatment for HIV impacting HSV.

genetics - Pedigree Probability of Autosomal Recessive Trait

Starting with the left hand side of the diagram:

• III:2 is definitely a carrier (Tt) as one parent (II:2) is affected (tt).


• III:1 is also definitely a carrier (Tt) as when mating with III:2 they produce an affected (tt) offspring (IV:1)


• This means that we can work out the possibilities for IV:4 as we know the parent genotypes. It follows the standard arrangement for two carrier parents giving the options of:
  
  • TT (1/4)
  
  
  • Tt (2/4 = 1/2)
  
  
  • tt (Normally 1/4 but in this case 0 as individual not marked as affected).
  
  


• Therefore for this scenario, the probabilities for IV:4 are :
  
  • TT (1/3)
  
  
  • Tt (2/3)
  
  

Now if we look at the right hand side of the diagram.

• IV:5 is definitely a carrier (Tt) as one of their parents (III:5) is affected.

This gives two possible Punnett squares to be examined:

|-------------------------------------------------------------------------------|
|                                        ♂ (IV:5)                               |
|                          T                                   t                |
|            -------------------------------------------------------------------|
|           |                                |                                  |
|         T |              TT                |                Tt                |
|           |                                |                                  |
| (IV:4)    |-------------------------------------------------------------------|
|    ♀      |                                |                                  |
|         T |              TT                |                Tt                |
|           |                                |                                  |
|-----------|-------------------------------------------------------------------|

This gives nil affected offspring so we can disregard this option for your question (as we are ONLY looking for scenarios which produce affected individuals).

Therefore the alternative is:

|-------------------------------------------------------------------------------|
|                                        ♂ (IV:5)                               |
|                          T                                   t                |
|            -------------------------------------------------------------------|
|           |                                |                                  |
|         T |              TT                |                Tt                |
|           |                                |                                  |
| (IV:4)    |-------------------------------------------------------------------|
|    ♀      |                                |                                  |
|         t |              TT                |                tt                |
|           |                                |                                  |
|-----------|-------------------------------------------------------------------|

Giving 1/4 affected offspring.

As mentioned above, in order to have affected offspring then IV:4 must be Tt. There is a 2/3 chance of this being the case. If this is the case, then there is a 1/4 chance of the child being tt.

Both conditions need to be true for this to happen so we multiply the fractions:

2/3 * 1/4 = 1/6

molecular biology - PDB Mining: Why Do I Find Atoms Less than 1 Angstrom Apart?

I am attempting to find potential Hydrogen bonds between Hydrogen donors and aromatic ring acceptors. I do this by predicting the location of Hydrogens on residues and then calculating how far these Hydrogens are from aromatic rings. If a certain Hydrogen is <7.0 Angstroms from a certain aromatic ring, then I take it under consideration: I form the N-H vector, which is the vector created by the Hydrogen under question and the Nitrogen in the backbone of the residue that the Hydrogen belongs to. I test that this N-H vector is pointing toward the plane of the aromatic ring, and I also test that the point of intersection between the plane of the aromatic and the N-H vector is within 6 Angstroms of the center of the aromatic ring.

If all of these conditions are met, then I consider it a Hydrogen bond between the Hydrogen and the aromatic ring. However, my data must be incorrect, because I am seeing situations where a Hydrogen is < 1.0 Angstrom from the plane of the aromatic. Atoms should not be getting that close to each other.

I thoroughly tested my method by hand using an example situation where my code identified one of the sidechain Hydrogens on an ASN is 0.3 Angstroms from the plane of the aromatic of a TRP. Unfortunately, I could not find any bugs. You can find a PDF of this verification here.

Any suggestions on how my method might be flawed would be greatly appreciated.

terminology - Calculating Protein Concentration from Kilo Units (KU)

The figure of 350 - 600 Units per mg refers to the specific activity of the enzyme.

The Unit is International Unit or IU and is usually defined as that amount of enzyme that will catalyze the transformation of 1 micromole of substrate (or product) per min, under defined assay conditions (such as pH, temperature, substrate concentration, presence of Mg⁺⁺, etc). It is thus a measure of activity.

When the enzyme is pure (no other extraeneous proteins present), the specific activity provides important information about the catalytic capacity of the enzyme.

It is usually calculated by measuring

• the activity of the enzyme preparation under defined assay conditions


• the protein concentration of the same enzyme preparation (using, say, the Lowry or Biuret method for protein estimation).

  Alternatively, if the E(1%, 280) is known (see below) and the enzyme is pure, measurement of the absorbance at 280 nm gives a very good estimate of protein content (and the enzyme may be recovered 'unharmed' at the end of the measurement).

Thus, taking a figure of 450 Units/mg for the specific activity of pyruvate kinase,
25 KU (25 Kilo-Units, I presume) contains 500/9 mg (~55 mg) protein.

I notice that the Sigma product sheet provides a figure for E(0.1%, 280) = 0.54.

This means that a 1 mg/ml solution of the protein will have an absorbance at 280 nm of 0.54

E(0.1%, 280) can be used as a very convenient measure of the protein content provided that the enzyme preparation supplied by Sigma is pure.

A 'rule of thumb', useful when the E(0.1%, 280) is unknown, is that a 1mg/ml protein solution has an A₂₈₀ of 1.

Thus if, say, the A₂₈₀ (absorbance at 280 nm) of the resuspended lyophilized powder is 1.08 and you have 5 ml of this, the protein concentration is 2mg/ml and you have 10 mg of protein in total. You may wish to assay the enzyme yourself to determine an accurate specific activity.

The EC (Enzyme Commission) Number may also be of interest. For pyruvate kinase (EC 2.7.1.40) see here.

For a great ref on PK (pdf may be downloaded) see here (Ainsworth et al.)

---

For your second question, I do not have access to that paper from home.

However, if calmodulin has a specific activity of 40 000 Units/mg,

• 25 000 Units is equivalent to 0.625 mg; this is in a volume of
  0.5 ml. Therefore, the calmodulin concentration is 1.25mg/ml.



• Taking the molecular weight of calmodulin to be 16 000,
  then 16 000 mg /ml (theoretical) would be a 1 Molar solution.
  Thus a 1.25 mg/ml solution is about 78 micromolar.

immunology - Harmless virus? - Biology

It is possible for viruses to live in mutualistic relationships with their hosts, these associations are often overlooked due to the devastating effect that many viruses can have.

To give an example in humans, when HIV-1-infected patients are also infected with hepatitis G virus, progression to AIDS is slowed significantly (Heringlake et al., 1998; Tillmann et al., 2001). Also hepatitis A infection can surpress hepatitis C infection (Deterding et al., 2006).

There are many other notable examples within plants, fungi, insects, and other animals, reviewed by Shen (2009), and Roossinck (2011), in two excellent papers.

The table below, summarises some beneficial viruses across all organisms, and is taken from Roossinck (2011).

References

• Deterding, K. et al., 2006. Hepatitis A virus infection suppresses hepatitis C virus replication and may lead to clearance of HCV. Journal of Hepatology, 45(6), pp.770-778.


• Heringlake, S. et al., 1998. GB Virus C/Hepatitis G Virus Infection: A Favorable Prognostic Factor in Human Immunodeficiency Virus-Infected Patients? Journal of Infectious Diseases, 177(6), pp.1723 -1726.


• Roossinck, M.J., 2011. The good viruses: viral mutualistic symbioses. Nature Reviews Microbiology, 9(2), pp.99–108.


• Shen, H.-H., 2009. The challenge of discovering beneficial viruses. Journal of Medical Microbiology, 58(4), pp.531 -532.


• Tillmann, H.L. et al., 2001. Infection with GB Virus C and Reduced Mortality among HIV-Infected Patients. New England Journal of Medicine, 345(10), pp.715-724.

genetics - T7 promoter leakiness

Can a gene be expressed under the T7 promoter in an E. coli strain (e.g. DH5 alpha), which does not have the T7 polymerase gene encoded in its genome? In other words, is T7 promoter leaky?

To be more specific, how is it possible that a regular E. coli strain, which does not encode for the T7 polymerase, can grow on kan selective media if it was transformed with a plasmid that has the kanR gene under T7 promoter?

structural biology - How many human proteins are very well characterized?

To address your list:

• a high quality 3D structure: this you can easily get from PDB, using the answers to the question you linked as starting point. However, it is become increasingly clear that intrinsically unstructured proteins also play important roles in the cell, and for these you won't get a good 3D structure.


• known activity in vivo / known associates in its activities in the cell: interaction partners can be found in databases like STRING, although just knowing the partners doesn't mean we understand what's going on :)


• kinetics of its activity: I know of no good resource on this one except BRENDA for enzyme kinetics. But of course there are also kinetics to protein–protein interactions that would be interesting to know. (Update: There is AffinDB.)


• regulatory elements for expression: there are databases like TransFac, but ChIP-Seq experiments show that we only know a tiny part of the regulation.

But, I think the key to a good answer lies here:

(probably most importantly) a general consensus among experts that it is well described

I would check how often the genes are described in the literature. The GeneRIFs from NCBI may be a good starting point, because these are snippets from the literature that are curated and shown on the NCBI Gene pages. E.g. p53 has >3800 of these. The length of the summary could also be a good proxy. (Or, the length of the Wikipedia page?)

evolution - Features in individuals causing high population variation

As I understand it, a population with high variation is something sought after, since it makes the population better equipped to face a dynamic environment.

Then, I guess features in an individual which causes it's population's variation to be high, should be selected for (high mutation rate, other features.. ?).

If so, does this manifest itself partly as individuals preferring to mate with individuals from another population? (With genetic difference within reason, perhaps a norm for a proffered genetic difference between individuals)

metabolism - How does the human body metabolize gasoline?

Gasoline toxicity through ingestions seems to be a topic where there's not a great deal of in-depth information available. I don't know how this works for chronic use, as most literature refers to acute scenarios. Either way, orally ingested, 30-50g is said to be toxic to humans while 350g can be fatal.[3].

So...

Gasoline's Constituents

A lot of components that make up gasoline are toxic to humans. This includes for example, benzene, toluene, xylene and butadiene. It's a mixture of more than 500 hydrocarbons and additives made up of:

• 60–70% alkanes (paraffins)


• 25–30% aromatics


• 6–9% alkenes (olefins).[2].

If you really want to know specifics about metabolism of gasoline, you can probably check out how some of the constituents are metabolised (processed) and their effects. This is because different components have varying metabolic pathways.

If you want to check this out, see Reese, et al at http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1520023/pdf/envhper00383-0118.pdf.[3].

Ingestion and Toxicity

Apparently, most reported cases of toxicity from gasoline occur from inhalation or absorption through the skin (intravenous use has also been reported). Even though ingestion is a frequent occurrence, there's not much data on outcomes after oral ingestion.[1],[2]. Gasoline can however be well absorbed through the gastrointestinal tract[2].

The main target organ of gasoline toxicity is the nervous system and at high doses, this effect can cause death within minutes.[3]. However, generally, the primary cause of mortality seems to be related to gasoline's toxicity to the lungs. There are severe effects on the pulmonary system. Other effects of ingestion hasn't been as well documented.[1],[2].

It's suggested that these compounds have a direct effect on lung tissue and disrupts gas exchange and causes fluid buildup in the lungs (pulmonary edema). This in turn causes the oxygen levels in the body to drop (hypoxemia).[2]. There are many other toxic effects on the lungs as well.

Additionally, liver damage, kidney damage, damage to blood cells, gastric ulcers and toxicity in the heart can occur.[2],[3]. Again, this is discussed in Reese, et al.

Hope that helps!

1. Rahman I. Gasoline ingestion: a rare cause of pancytopenia. Am J Med Sci. 2009. 338(5):433-4.


2. Domej W. Successful outcome after intravenous gasoline injection. J Med Toxicol. 2007. 3(4):173-7.


3. Reese, et al. Acute Toxicity of Gasoline and Some Additives. Environmental Health Perspectives. 1993. 115-131.

dna - How does one measure the length of telomeres?

Again, not an expert, but I found this article seems to say it pretty well:

Telomere's are most commonly measured by qPCR of the repetitive regions with degenerate oligomer sequences such as "TTAGGG and CCCTAA repeats" Although articles like this one appear to advance measurements in vertebrates, this reference from 2011 implies two techniques continue to be most common:

1) qPCR with the appropriate oligos and then running them on an agarose gel

2) Southern blot of a restriction digest that leaves the telomeres intact

Clearly sequencing techniques don't do well with repetitive sequences such as this, so in general this still seems like the most common method.

This would indeed be quite destructive, at least for the cells used, though a biopsy would be enough, though apparently micrograms of DNA are required. Kind of a large amount.

The accuracy would then be limited to that of the gel or blot. That would generally be a percentage of total length, I think 2% might be obtainable, but I'm not sure about that part.

human biology - Why do we age? or Do we have a theory of senescence?

There is a pretty good discussion on this topic in chapter 2 of Geriactric Medicine - An Evidence Based Approach (4th ed) by Cassel. This is the main reference for the info below which can hopefully add something to the answers already given.

In terms of views on ageing, there's evidence to support both:

• general principles that may apply to it; and


• it being a consequence of a collection of degenerative processes (this is apparently the more supported view).

Since almost all biological systems in the body degenerates with age and this happens seemingly at random, it's been difficult to identify particular catalysts that cause this. Consequently, biologists apparently steer away from a general theory or mechanism.

However, there are two classes of theories that have been floating around. That is 'loose cannon' and 'weak link'.

Loose Cannon encompasses theories that support the 'wear and tear' proposition. Two popular theories under this banner are free radicals and glucose.

Weak Link suggests that particular physiological systems are vulnerable during senescence and if a system fails, the whole body begins to decline. It's suggested that the neuroendocrine and immune systems are particularly vulnerable.

There is also a limit on the ability of cells to replicate - this is called the Hayflick Phenomenon (or limit). The reduction of the enzyme Telomerase, which lengthens telomeres during mitosis, is implicated in limiting a cell's ability to replicate indefinitely.

zoology - Could an animal with an open circulatory system survive in a near-zero gravity environment?

An animal such as a crayfish relies on gravity to keep its circulatory system running. If it is turned upside down, the gravity works against the system, suffocating the animal. Now if, instead of placing the animal on it's back, we were to take take it to a space station, gravity would not be working against it, and random blood flow plus capillary action should get some blood flowing. Would the animal survive this?

molecular biology - Does the MS2 RNA binding protein have any translational repression effects?

I assume you are talking about the mTAG visualization technique for mRNAs (1). You are probably familiar with it, but the OFR of interest is tagged with MS2L sequence downstream of it and upstream of the 3'UTR. There is also a modified version, in which you can visualize both the mRNA of interest, and the protein that is translated, see the figure below (2). Haim and colleagues found that yeast expressing mCherry and MS2L tagged ATP2 grew on glycerol-containing media, which would require efficient translation of ATP2.

So I would assume that there is no translational repression if you tag the mRNA downstream of the ORF. Unless you have a very good reason to tag the ORF of interest with MS2 upstream of the start codon, I wouldn't do it, because these stem-loop structures would most probably stall translation, as mRNA secondary structure has been implicated in translation initiation efficiency.

botany - How do trees manage to grow equally in all directions?

There are some other good answers which provide part of the picture, but I think there is a fundamental organising principle which has been missed. Konrad has touched on it in his answer.

The reason trees, and most plants, tend to grow equally in all directions is that they have iteratively generated branching and radial symmetry which is controlled in a feedback loop of the growth promoting hormone auxin and auxin-sensitive auxin transporters. This is an elegant biological algorithm which explains all branching growth.

The things Konrad identifies (phototropism, gravitropism, etc.) serve as orientation cues which help the plant determine which axes to grow along, but fundamentally the process is about auxin gradients. There are exceptions, as others have pointed out in their answers, and they usually result from severe imbalances in the orientation cues.

I'll try to explain the growth process clearly (and it gives me an opportunity to try my hand at diagramming again ^_^)...

---

Auxin is a plant hormone (actually a class of hormones, but mostly when people say auxin, they mean indole-3-acetic acid) which promotes cell elongation and division. The basic principle which allows auxin to act in the organising way it does is that auxin is produced inside cells, and proteins which export auxin from a cell develop on the side of the cell which has the highest auxin concentration (see figure below).

So auxin gets transported up the concentration gradient of auxin! Thus if you get an area of high auxin concentration developing somehow, more auxin is then transported towards that area. An area of high auxin concentration relative to the surrounding tissue is called an auxin maximum (plural 'maxima').

For most of the life of the plant, auxin is produced pretty much equally in most cells. However, at the very early stages of embryo development, it gets produced preferentially along the embryonic axis (see figure below, part 1). That creates a meristem - a group of cells where cell division is taking place - at the auxin maximum at each end of the embryo. Since this particular meristem is at the apex of the plant, it is called the apical meristem, and it is usually the strongest one in the plant.

So by having a meristem at each end, the embryo then elongates as cell division is only taking place at those points. This leads to part 2 of the image above, where the two meristems get so far apart that the auxin gradient is so weak as to no longer have its organising effect (area in the red square). When that happens, the auxin produced in cells in that area concentrates in a chaotic way for a short time until another center of transport is created. This happens, as the first one did, when a particular area of the tissue has a slightly higher concentration of auxin, and so auxin in the surrounding tissue is transported towards it. This leads to part 3 of the figure, in which two new meristems are created on the sides of the plant (called lateral meristems).

Lateral meristems are where branches occur on plants. If you then imagine this process continuing to iterate over and over, you will see that the branches, as they elongate, will develop meristems at the tips and along the sides. The main stem will also continue elongating, and develop more lateral stems. The root will begin to branch, and those branches will branch, etc. If you can understand how this elegant system works, you understand how plants grow, and why they grow in repeating units as opposed to in a body plan like animals.

It also explains why, if you cut off the tip of a stem, it promotes branching. By removing the apical meristem, you get rid of the auxin gradient and enable the creating of multiple smaller meristems which each develop into branches.

So far I've explained regular branching, but the same system causes the radial symmetry which makes trees (usually) grow in all directions equally...

Imagine taking a cross section through a stem and looking down all the way through it (as depicted crudely above). Just as auxin gradients act to coordinate growth along the length of the plant, they also coordinate it radially, as the maxima will tend to space themselves out as far from one another as possible. That leads to branches growing in all directions equally (on average).

I welcome comments on this answer, as I think its so important to understanding plant growth that I'd like to hone my answer to make it as good as possible.

homework - What is the main general difference between Mitosis and Meiosis?

I found such a clause:

The general principle is that mitosis creates somatic cells and
meiosis creates germ cells.

However, I cannot agree. Each gametogonium needs to go through mitosis before it can enter meiosis I. So in that case mitosis is happening with germ cells so the clause is false.

I would rephrase the sentence to be

The general principle is that meiosis creates only germ cells with the possibility of a decrease in chromosome number, while mitosis can create both somatic and germ cells while the ploidy stays constant.

Ok, not perfect.

How would you say the main general difference between mitosis and meiosis?

Pipetting damage on cells - Biology

Shear stress $tau$ in this small sizes is usually measured in dyne/cm2 or N/m2 = Pa. The equations betweeen them: $1dyn/cm^2 = 10^{-5}N/cm^2 = 0.1N/m^2 = 0.1Pa$.

What kind of damages zygotes can suffer by pipetting?

Using scanning electron microscopy, we found open holes on the surface
of lysed eggs, indicating failure of the plasma membrane to reseal
after microinjection. No holes were seen in unlysed eggs, but many of
them had membrane alterations suggestive of healed punctures.

Even a small 1.2 dyn/cm2 shear stress induces apoptosis by pipetting zygotes. So zygotes have their critical shear stress level by 1.2 dyn/cm2 and pipetting involves greater forces than 1.2 dyn/cm2.

Shear stress at 1.2 dynes/cm2 induces stress-activated protein
kinase/jun kinase phosphorylation that precedes and causes apoptosis
in embryos (Xie et al., 2006b, Biol Reprod). Pipetting embryos is
necessary for many protocols, from in vitro fertilization to
collecting embryos prior to analyzing gene expression by microarrays.
We sought to determine if pipetting upregulates phosphorylated MAPK8/9
(formerly known as stress-activated protein kinase/jun
kinase/SAPK/JNK1, 2). We found that phosphorylated MAPK8/9, a marker
of MAPK8/9 activation, is upregulated in a dose-dependent manner by
pipetting.

The critical shear stress level is somewhere between 0.01 and 1000 dyn/cm2 by animal cells depending on the cell type and species. (I think the average is somewhere about 50 dyn/cm2, but it is very hard to differentiate between articles mentioning critical shear levels and most lethal shear levels, so the range and the average might be lower.) The death constant (1/h) increases exponentially by increasing the shear stress.

An apparatus for the detailed investigation of the influence of shear
stress on adherent BHK cells was developed. Shear forces between 0.0
and 2.5 N m−2 were studied. The influence on cell viability, cell
morphology, cell lysis, and cell size was determined. Increasing shear
forces as well as increasing exposure duration caused increasing
changes in cell morphology and cell death. A “critical shear stress
level” was determined.

Shear stress related damage to a mouse hybridoma was examined by
Abu-Reesh and Kargi under laminar and turbulent conditions in a
coaxial cylinder Searle viscometer. Cells were exposed to 5 to 100
N/m2 shear stress levels for 0.5 to 3.0 h. At a given shear stress and
exposure time, turbulent shear was much more damaging than laminar
shear as also reported in the past for protozoa and plant cells. Under
turbulent conditions, damage occurred when shear stress exceeded 5
N/m2. Respiratory activity of the cells was damaged earlier than the
cell membrane, thus implying transmission of the stress signal to the
interior of the cell. Cell damage followed first-order kinetics both
in laminar and turbulent environments. For turbulent shear stress
levels of 5 to 30 N/m2, the death rate constant (kd) increased
exponentially with increasing stress level; the kd values varied over
0.1 to 1.0 1/h.

Subconfluent endothelial cultures continuously exposed to 1–5
dynes/cm2 shear proliferate at a rate comparable to that of static
cultures and reach the same saturation density (≃ 1.0–1.5 × 105
cells/cm2 ). When exposed to a laminar shear stress of 5–10 dynes/cm2
, confluent monolayers undergo a time-dependent change in cell shape
from polygonal to ellipsoidal and become uniformly oriented with flow.
Regeneration of linear “wounds” in confluent monolayer appears to be
influenced by the direction of the applied force. Preliminary studies
indicate that certain endothelial cell functions, including fluid
endocytosis, cytoskeletal assembly and nonthrombogenic surface
properties, also are sensitive to shear stress. These observations
suggest that fluid mechanical forces can directly influence
endothelial cell structure and function.

Shear stress above 0.25 dyne/cm(2) resulted in dramatic loss of
podocytes but not of proximal tubular epithelial cells (LLC-PK(1)
cells) after 20 h.

A series of careful studies has been made on blood damage in a
rotational viscometer. Specific attention has been focused on the
effects of solid surface interaction, centrifugal force, air interface
interaction, mixing of sheared and unsheared layers, cell-cell
interaction, and viscous heating. The results show that there is a
threshold shear stress, 1500 dynes/cm2, above which extensive cell
damage is directly due to shear stress, and the various secondary
effects listed above are negligible.

The shear stress threshold of some dinoflagellates (microalgae) is
even lower than that of erythrocytes (0.029 N/m2). For example, a
continuous laminar shear stress level of only 0.0044 N/m2 (equivalent
to a shear rate of 2.2 1/s) has proved lethal to the dinoflagellate
Gonyaulax polyedra.

Other cell types are not necessary as sensitive as animal cells and they don't necessary react with apoptosis (about 10 dyn/cm2) to shear stress, so you have to use necrotic (about 5000 dyn/cm2) forces to destroy them :

cell type                       size                shear sensitivity
microbial cells                 1-10μm              low
microbial pellets/clumps        up to 1cm           moderate
plant cells                     100μm               moderate/high
plant cell aggregates           up to 1-2cm         high
animal cells                    20μm                high
animal cells on microcarriers   80-200μm            very high
fungi cells                     2-10μm              moderate/high

Results show that Chinese Hamster Ovaries and Human Embryonic Kidney
cells will enter the apoptotic pathway when subjected to low levels of
hydrodynamic stress (around 2.0 Pa) in oscillating, extensional flow.
In contrast, necrotic death prevails when the cells are exposed to
hydrodynamic stresses around 1.0 Pa in simple shear flow or around
500 Pa in extensional flow.

The shear sensitivity is not determined only by cell type and species, there are many other factors involved:

• type of cell and species


• composition and thickness of cell wall when present


• size and morphology of cell


• the intensity and nature of shear stress, whether turbulent or laminar, or associated with interfaces (e.g. during bubble rise and rupture)


• growth history, both short-term (e.g. starvation) and long term adaptation


• growth medium (trace elements, vitamins, carbon and nitrogen sources)


• growth rate


• growth stage


• type and concentration of shear protective agents if present

Cells can be very sensitive to shear stress caused by turbulent flow, while not so sensitive to shear stress caused by laminar flow.

On the basis of laminar flow viscometriy measurements, a critical
shear stress level of 80-200 N/m2 has been suggested for Morindata
citrifolia cells.

... while for Daucus carota a shear stress level of 50 N/m2 has been
associated with cell damage. In other study, carrot cells in a laminar
flow Couette viscosimeter lost the ability to grow and divide in the
shear stress range of 0.5-100 N/m2. The intracellular enzyme activity
was impaired at shear stress levels above 3000N/m2, but significant
lysis did not occur until a shear stress level of 10.000 N/m2 applied
over a prolonged perioud (>1h).

In contrast to the behavior in laminar flow, the cells were quite
sensitive to turbulent impeller agitation. Impeller tip speeds of ~1.1
m/s lysed a significant proportion of the cells within 40min.

The bubble damage is severe (1000 cells by a single 3.5mm size bubble) because of the cell adherence to the interface of the bubble and the strong forces involved (>1000 dyn/cm2 by stirred bioreactors). The adhesion and so the damage can be reduced with surfactants.

It is proposed that when cells are either attached to, or very near, a
rupturing bubble, the hydrodynamic forces associated with the rupture
are sufficient to kill the cells.

All experiments were conducted with Spodoptera frugiperda (SF-9)
insect cells, in TNM-FH and SFML medium, with and without Pluronic
F-68. Experiments indicate that approximately 1050 cells are killed
per single, 3.5-mm bubble rupture in TNM-FH medium and approximately
the same number of dead cells are present in the upward jet. It was
also observed that the concentration of cells in this upward jet is
higher than the cell suspension in TNM-FH medium without Pluronic F-68
by a factor of two. It is believed that this higher concentration is
the result of cells adhering to the bubble interface. These cells are
swept up into the upward jet during the bubble rupture process.
Finally, it is suggested that a thin layer around the bubble
containing these absorbed cells is the “hypothetical killing volume”
presented by other researchers.

For a hybridoma line, reported that exposure to laminar shear stress
(208 N/m2) in unaerated flow in a cone and plate viscometer led to
substantial loss in cell count and viability within 20 min. At a
constant 180 s exposure, increasing shear stress over 100-350 N/m2
linearly enhanced cell disruption, with >90% of the cells being
destroyed at 350 N/m2 stress level. Shear stres levels associated with
bubble rupture at the surface of a bioreactor may range over 100-300
N/m2. These values are remarkably consistent with shear rates that
damaged hybridomas in unaerated laminar flow experiments.

Smaller hole pipettes cause more damage.

We also examined aspects of the gene transfer procedure that might
influence survival such as the size of injection pipettes and their
taper relative to zygote diameter, possible toxicity of the injection
medium, the timing of injection, and immediate vs. delayed pipette
withdrawal. The only factors that significantly affected cell
viability were pipette size and taper, and timing of injection in
relation to first cleavage. This suggests that zygote viability
correlates inversely with the size of the hole produced by the
injection pipette and that damage to the membrane is less successfully
repaired as the fertilized egg readies itself for division.

It is hard to find anything about the level of shear stress by pipetting. It can be certainly more than 1 dyn/cm2. It has a short duration (at most a few seconds). I think the following factors can influence the shear stress levels by pipetting:

• pipette type


• flow speed (faster flow can be more likely turbulent)
  


• bubble formation

Probably more factors are involved but I am not a pipetting expert. ;-) I agree with the others, it surely depends on the personal skills e.g. an amateur can create huge bubbles by pipetting, which can kill a lot of cells by formation and disruption...

I agree with Artem that this is an experiment to do especially if the result is important for you. What you need to create a model about pipette damage, are the shear stress levels by pipetting and the critical shear stress levels of the cells. I think it is hard to design and experiment in which you can measure the shear stress levels in your pipettes and there is no flow model for pipetting as far as I know, so it can be a good topic for a thesis or a diploma work.

microbiology - Are single-celled organisms capable of learning?

I'd like to know what is the reference for amoebic learning. I cannot comment directly on this, but there is some evidence for "adaptive anticipation" in both prokaryotes and single-celled Eukaryotes which do not have a nervous system.

In the case of E. coli, it has been shown that the bacteria can anticipate the environment it is about to enter. E. coli in the digestive tracts of mammals will typically be exposed to initially a lactose, and then later to a maltose environment as the bacteria pass down through the animal tract. This suggests that upon encountering a lactose environment, maltose operons are induced. I.e., upon encountering lactose, maltose is anticipated. This suggests a "genetic memory" of the sequence of sugar types where lactose is always encountered before maltose.

Further cultures (500 generations) of E. coli in the absence of maltose but in the presence of lactose reduced the maltose operon activity to negligible levels, suggesting that this is an adaptive prediction of environmental changes.

Mitchell, A et al., Adaptive Prediction of environmental changes by microorganisms, 2009, 460, 1038

Are there any examples of sudden leaps in evolution?

@kmm and @shigeta provided you with a nice observational account of sudden leaps in large organisms. However, if you want to look at where this is the norm and try to build a mathematical theory then you need to look at something much smaller; the prime candidate is affinity maturation.

In the human immune system, when exposed to an antigen B cells produce antibodies. If it is your first exposure to the antigen then the antibodies produced will probably have very low binding affinity. However, after some exposure time, your B cells will start to produce antibodies with much higher affinities for the antigen and thus you will be able to better fight off the disease. The cool part, is that the antigen produced is tune via an evolutionary process!

There is differential survival, with only antibodies with the highest affinity being able to survive. Variability is introduced by a very high mutation rate in the complementarity determing region (CDR). (Tonegawa, 1983). The length of this evolutionary process is very short, typically a local equilibrium is found after only 6-8 nucleotide changes in CDR (Crews et al., 1981; Tonegawa, 1983; Clark et al., 1985), so you need only a few point mutations to quickly develop a drastically better tuned antibody.

The standard mathematical model for this is Kauffman's NK model. With a protein sequence on $N$ sites, we say that evolution is fast (and we have a sudden leap) if after our fitness landscape changes, we can get to a new local equilibrium in a number of generations that scales with $log N$. Kauffman & Weinberger (1989) showed how this model can be used to study affinity maturation, and showed that to achieve a sudden leap we need high epistasis and low correlations between pointwise mutants. In particular, their model suggests that typical epistasis in the CDR is on the order of 40 proteins (out of the total 112 proteins in the CDR).

---

References

Clark, S.H., Huppi, K., Ruezinsky, D., Staudt, L., Gerhard, W., & Weigert, M. (1985). Inter- and intraclonal diversity in the antibody response to influenza hemagglutin. J. Exp. Med. 161, 687.

Crews, S., Griffin, J., Huang, H., Calame, K., & Hood, L. (1981). A single V gene segment encodes the immune response to phosphorylcholine: somatic mutation is correlated with the class of the antibody. Cell 25, 59.

Kauffman, S. and Weinberger, E. (1989) The NK Model of rugged fitness landscapes and its application to the maturation of the immune response. Journal of Theoretical Biology, 141(2): 211-245

Tonegawa, S. (1983). Somatic generation of antibody diversity. Nature 302, 575.

bioinorganic chemistry - What is the molecular mechanism of cystine bond formation?

If your question is with respect to a eukaryotic cell, the di-sulfide bridge/bond is formed in the rough endoplasmic reticulum which is an oxidative environment (unlike most other organelles which are reductive). This paper may be of relevance to you:

Pathways for protein disulphide bond formation - Frand et al, Trends Cell Biol. 2000 May;10(5):203-10.

human biology - What causes knuckle "popping" and the feeling of relief that comes from it?

Nobody really knows where it comes from. The currently most popular theory is that pulling the joint apart leads the gases in the joint's cartilage to accumulate and form a bubble which then pops when you let it spring back. The only thing that has been researched is whether it has an effect on the joint, but people who do it regularly don't seem to have any problems with their joints more than anyone else (I'll find the reference once I have time). Apart from that it's hard to get funding for this kind of research because it has little practical use.

Relief like feeling? That must be subjective to you, I used to do it every now and then but I don't remember getting any relief from it. In that case, it's probably the same relief as other people get from picking their nose, their ears, biting their nails, etc. It's called compensation reaction.

human biology - What causes fingerprints to form and why is the pattern formed unique?

I would say genetic diversity is the primary reason which results in other reasons that you are looking for. At the lowest level, random crossing over at prophase I, random separation of homologous chromosomes at anaphase I, random separation of sister chromatids at anaphase II, and random fertilization: one sperm fertilizes one egg randomly.

The skin is developed from ectoderm so need to look at the formation of embryonic disc and specifically to the genesis of germ layers: ectoderm.

However, I would stick to the primary reasons, since it is extremely difficult to visualize the given formation - actually we do not have resources for it at the moment.

---

Very good question the last part. I have an intuition that skin develops randomly because of the above reasons. You would also need a lot of memory to make identical skins for twins! It has not been useful to have identical fingerprints between two people so evolution has not resulted into it.

Feeling surfaces and gripping are movements - not much space taken things, in contrast to the memory needed in storing the exact surfaces of skin from one generation to another. - Learning is a way to save resources here and it is a lot more efficient and than storing static information to species from one generation to another.

genetics - How many gigabytes of DNA are there on earth?

If you simply take one order of insects, Coleoptera, there are just under 400,000 described species with estimates from 850,000 to 4,000,000 species total in just this order. The number of primates is under 1,000. If your assumption of say 10MB for all other primates would be accurate, just adding in the low end estimate of 850,000 at 10MB per 1000 we are quickly at 8,500GB which seems to be a factorial out of the GB range.

So, we have a broad estimate of non-bacterial of plants, animals etc. at say 8,700,000.

Jason Gans found in a 1 gram of soil survey approximatly 1,000,000 bacterial species.

SO the total accounting for species number is totally impossible to estimate for anything at this time, let alone the genome.

Even for something as "common" as a giraffe, there are up to 9 sub-species with genome differences within each subspecies.

So, once we get them all decribed, we can then work on the genome sequence for each and get you some answers!

Cell proliferation limit and senescence of embryonic stem cells and fibroblasts

Telomeres (caps on the ends of chromosomes that are gradually shortened during each cell division) determine the maximum number of times a cell can divide, known as the 'Hayflick limit'. Cells that are able to express telomerase, an enzyme capable of extending the telomeres, can divide indefinitely.

There are various types of stem cell in a body which replenish the 'pool' of cells in the tissue once they reach the end of their useful lifespan. Each stem cell expresses telomerase to lengthen the telomeres after a round of cell division. The inhibition of telomerase in somatic (normal) cells is a major 'challenge' for cancerous cells - in order to achieve uninhibited proliferation, they must express telomerase. Thus, there are many molecular similarities between stem cells and cancerous cells (well, with regards to telomerase, anyway!) [ref].

Fibroblasts are somatic cells - they are the 'normal' cells that make up the vast majority of organsisms. The only cells not known as somatic are gametes/germ cells and stem cells. They thus do not express telomerase, and have a replicative lifespan, also known as a Hayflick limit.

With regard to your question about engineering organs, there are many resident stem cells in tissues (and thus organs) that 'top-up' the somatic cells when required. As organisms age, this pool of stem cells becomes depleted [ref]. Artificially engineering an organ such as the heart (I assume this is what you mean) would be inherently difficult, as many vascular cells are not replaced, but can last a whole lifetime - this is also true for many neuronal cells. So replicative lifespan is not necessarily related to lifespan!

Does extracted DNA degrade after a certain time period?

For direct use as template in PCR runs. Chelex 100 5-10% w/v extraction. Without listing the whole protocol, in the end the supernate is decanted off and then stored at 4°C. I was under the impression that this could be stored and later used almost indefinately but two of four samples extracted several months back failed to produce a product (when it was known they should have).

Assuming no mistakes were made and the reactions were the same, is there a technical reason the template would degrade to an unusable point?

Is there a rule of thumb about how long it can reasonably be expected to last?

neuroscience - Brain + ethanol experiment suggestions needed

The main problem with what you are asking is that you want to show effects on vigilance and memory in vitro. That is just not possible: if you want vigilance and memory you need a live animal, there is no way around it.

Next point: you have an audience of non scientists, so you can lose them very quickly if you start speaking about NMDA, LTP or similar things without a clear explanation, so start with that: get a nice review (Pubmed is a good start, as always), and give a simple theoretical explanation of one of the mechanisms involved. It is then important to clearly explain the experiment you will perform, and what you are expecting to see in case there is an effect.

Unfortunately the best way to see effects on NMDA receptors would be electrophysiology. This is not trivial and, if you have never done it, I would not suggest going that way. However, if you have access to an electrophysiology setup you will most likely have access to someone who uses it, and that could make the experiment for you.

I am not an expert on this topic, but a quick research on Pubmed seems to indicate that alcohol can, for instance, reduce LTP in hippocampal neuron.
Maybe what you could do is having someone patch hippocampal neurons and show how an LTP protocol is used. You don't necessary have to show them the whole thing (I doubt anyone in a right state of mind would bare to stay until the whole experiment is performed in control condition first, and alcohol later...).

Also, the results of these experiments are often non obvious to analyse, so you should have some data prepared and analysed earlier and quickly show how analysis is performed, before showing the final results.

Calcium imaging experiments could also be used and would probably give results that are easier to understand for a laymen audience.

See for instance Fig. 1 or Fig. 5 of this paper (just the first one I found, there surely are other):
Ethanol alters calcium signaling in axonal growth cones.

homework - What is the structure and function of chromosomes during interphase?

So the DNA in some chromosomes must have the pieces of information about how to do the DNA replication. - I am not sure about thing.

Genomes contain what is called the "origin of replication" - specific sequences in the DNA that tell DNA polymerase where to bind and to initiate replication.

As for your main question, I'm a little confused as to what you're asking. In a general sense, chromosomes function as carriers of genetic information. In eukaryotes, nuclear DNA is organized in the nucleus on linear chromosomes which carry most of the genetic information an organism needs to survive. In bacteria, the chromosome is a circular piece of DNA. In eukaryotes, the chromosome is also bound by histone proteins, which serves to regulate expression of certain genes and to help anchor the chromosomes to the inner nuclear membrane.

genetics - What exactly is meant by the expression "differentially expressed"?

Although each cell of your body essentially contains the same DNA and the same genes, cells in different tissues express (turn on) different genes under different conditions. Measuring differential gene expression involves looking at the amount of expression for a gene (or set of genes) in two contrasting scenarios. The contrast could be across different times, different tissues, different conditions, different related species, etc.

When, you say a gene is "differentially expressed", this is very context-specific. The phrase means nothing by itself, and it is only useful in terms of the applicable contrast. For example, the statement "gene A is differentially expressed" is uninformative, while the statement "gene A is differentially expressed in liver and muscle tissue" is descriptive--it tells you that liver tissues and muscle tissues have a significantly different level of gene A products. Often the terms "up-regulated" and "down-regulated" are also used to provide additional detail. In the context of the previous example, the statement "gene A is up-regulated in muscle tissues" tells you that the level of gene A products is higher in muscle tissues than in liver tissues.

biochemistry - Can elements of one's environment act directly as hormones?

A hormone is defined as "a chemical released by a cell or a gland in one part of the body that sends out messages that affect cells in other parts of the organism" (I'm just taking Wikipedia definition).

Hormones work by binding to specific receptors present on their target cells so, if there is something in the environment that mimics the hormone, by binding to the same receptor they can act as hormones: these substances are called xenohormones and can often act as endocrine disruptors compounds (EDC), by acting on various organs in the body.

Probably the most known xenohormones are xenoestrogens that are been studied as possibly harmful for human health (e.g. linked to breast cancer), and as an environmental hazards, as they can, for instance, cause reproductive problems in fish.

Xenohormones are not necessarily bad, though. Some analogs of human hormones are used in therapy, having being synthesised specifically, for instance, to have higher potency then their natural counterparts.

homework - What is the total number of rounds of cleavage during mammalian embryonic development?

It's not a totally answerable question, since some types of cells are going to divide more times than others. But for an estimate, take as a starting proposition that there are 1 trillion cells in the adult human body. [1] The average weight for a human is 62kg. [2] Average birth weight is about 3.4 kg. [3]

So that implies roughtly (3.4/62)* 1 trillion = 55 billion cells in a newborn.

You then take the log base 2 of 55 billion, which gives you the exponent you have to hang on 2 in order to get 55 billion, which is about 35. Then add one for that additional cell division to get from one to two cells == 36 divisions.

Of course I'm just using math, not biology, so your actual reality may vary. Certainly some cells will reproduce more often than others, maybe cells actually grow in mass instead of dividing (i.e., baby cells might have less mass than adult cells) so the baby-cell-count could be off, lots of possible sources of error.

[1] http://www.nichd.nih.gov/publications/pubs/fragileX/sub3.cfm

[2] http://en.wikipedia.org/wiki/Body_weight#Average_weight_around_the_world

[3] http://en.wikipedia.org/wiki/Infant#Weight

human biology - If body temperature is 37°C (98.6°F), why are most people more comfortable at around 21°C (70°F)?

This is due to the fact that skin is the interface where heat is lost.

Our body due to constant functioning, produces heat constantly as a by-product (due to exothermic reaction of ATP break mainly). The excess heat needs to be conducted away from the body, or it will cause a decrease in the body metabolism to prevent temperature rise.

Heat is lost mainly through the skin by:

1. Sweating - Through evaporation


2. Radiation - As heat waves (IR rays - That's why IR camera captures people at night)


3. Conduction - Directly through objects that touch skin


4. Convection - Through air circulation

When the ambient temperature rises, the heat lost through radiation, conduction, and convection drastically decreases. And often when the temperature is high, there is a accompanying rise in the relative humidity which decreases the heat loss through sweating (as the amount of water vapor is high in the atmosphere, the sweat does not evaporate, so no heat is lost).

So the heat which is not lost is felt as the "hot sensation". It relieves by stopping any activity, seeking shade or a cool place, etc... all of which increases the heat lost or decreases the heat produced.

---

You have to note that the temperature of skin is lower than the body temperature.

The Skin temperature is lower than the core body temperature for two reasons:

1. The skin acts as a medium through which the external temperature is measured - as such the skin temperature is at equilibrium with the external temperature. The brain regulates the core-body temperature in response to temperature measured through skin. If a person is exposed suddenly to cold environment, skin looses much of its heat in the form of radiation (radiation is direclty proportional to the temperature difference) this will cause perception of cold and the body starts shivering even though no actual heat loss has occured from the core body thermal load (only skin looses heat, not the body core). The brain anticipates that the core will loose its heat when exposed to such low temperature for prolonged periods and starts the warming mechanism before the actual cooling occurs such that the cooling is either prevented or minimized. This is called Anticipatory control and the temperature of skin being close to the ambient temperature within physiological limits is needed for this.




2. Skin is the medium (almost the only medium) through which the excessive heat produced by the body core during activity is expelled. The skin temperature is lower so that a constant gradient can be created between the body core and body surface to maintain the flow of heat. (Heat losses through urine and feces is minimal)

For more details see this question. All the answers in this question are good and will increase your understanding of what actually happens.

zoology - Color of the Fur coat of Polar bears

The fur coats of polar bears are in fact clear tubes - not white fur as is often believed. It is their skin that is black, to absorb the most warmth from the sun (and the clear tubes allow the majority of the light to pass through to the skin).

I recently watched the BBC's Frozen Planet TV series (with David Attenborough), where I heard the above answer. If you haven't already, I recommend watching it.

molecular biology - What is causing my problem with very low yields when isolating a 42kb yeast plasmid?

I've successfully used the Zymoprep kit, but only for smaller plasmids in 2-hybrid experiments. So their cell lysis enzyme/buffer system works well, but I'd guess that your 42kb plasmid is precipitating out with the genomic DNA.

The standard Qiagen midi columns are capable of capturing large constructs. This has worked wonders for me when I was purifying large BACmids from E. coli - and gets the cost down to $10/sample (it does require additional buffer, but that's a minor expense). It might be worth trying the Qiagen yeast DNA protocols for the midi kit and see if you can isolate your large plasmid?

Is Batesian Mimicry a form a parasitism? To what extent is the species with real defenses harmed by the defenceless species?

In my own opinion, I would not classify this as parasitism, as more unpalatable species are eaten with Batesian mimicry (and eating causes death). Parasitic organisms often do not kill the host, whereas in this case, it does. However, you are correct in stating that a unpalatable species with aposematic coloring is detrimented by the presence of a palatable Batesian mimic.

Predators often recognize the aposematic coloring of an unpalatable or dangerous species. They "train" themselves not to eat that particular prey, as it is detrimental. The presence of a Batesian mimic which is palatable or not harmful, in simple terms, "untrains" predators of the harmful nature of that particular aposematic coloring. Predators are "confused", and are thus more likely to eat preys with that aposematic coloring, and results in more death of the species being mimicked, while providing protection to the species that is mimicking.

As far as I'm concerned, within an ecosystem, this phenomenon happens with any Batesian mimicry to some degree. Factors that may influence this degree is the recognizability of the aposematic coloring, as well as the ability of a predator to recognize this coloring.

I hope this clears things up.

biochemistry - Solution based measurement of Solvent-Accessible Surface Area of macromolecules

I only know of one method, but here it is. You create a sphere the diameter of the VdW radius of water, and then 'roll' it along the surface. I know this as a Richards-Lee surface, wikipedia has another name for it.

This looks complicated, but its not. you move the probe sphere along the surface of the molecule in the XY plane until it just touches the vdW radius of the protein, keeping the center of the sphere as the surface, all the way around the molecule. If you like, you can color the surface by the charge of the position too, which is useful for discussing solvent interactions.

Then you translate along the z axis and do another contour until you run out of protein. Apparently jmol and other packages will do this for you.

Wikipedia references a more mathematical method LCPO, which I am not so familiar with.

Is this accurate? As usual with such calculations its more of a guess than an answer. You can do the calculation on any structure or any ensemble of structures (like NMR gives). It doesn't understand how the molecule might be flexible or dynamic. If you read up on your physical chemistry you see that proteins breathe and can allow diffusion into the core rather readily. If I recall right, you can get rather large molecules quenching heme flouresence in hemoglobin at room temperature.

If you are looking to dock 2 proteins, SAS might be more useful. Its an important piece of information, but not an ultimate answer. I'm afraid with proteins that doesn't happen so easily.

@bobthejoe asked about SAS for which no structure exists.
This is an extremely difficult thing to even guess at. The non helpful answer is that the surface of the protein goes as the cube root of the molecular weight of the protein.

By getting a solution of the protein and shooting it in a syhcrotron, you can get a mean radius of gyration pretty easily which will give you an ellipsoidal volume (and surface area) for a protein. Again most of the particulars would be lost and this could easily be off by 25% for an irregularly shaped protein. For a regular globular protein it might give an answer similar to the power law above.

I have seen physical chemistry experiments that look for changes in osmotic pressure when the salt concentration in a solution of the protein changes substantially (Adrian Parsegian's work at NIH in the late 80s).

I doubt you will find any of these answers useful as their mean error is going to be very large (20-200%) and also assumes the protein is soluable and amenable to the experimental conditions.

Solvent probes can help too. For instance exposing the protein to D20 then doing mass spectroscopy on the protein. This is still only going to give you a general idea of how much of the peptide is surface exposed. Protein structure is still pretty necessary to getting any accurate measurement of SAS I think.

homework - Do humans have Coelom?

Kimball Biology 5e says

Coelom is the main body cavity of many animals. It is lined with an epithelium derived from mesoderm.

Gilbert Embryonal Biology 9e says

Coelom is the space between the somatic mesoderm and splanchnic mesoderm that becomes the body cavity. In mammals, the coelom becomes subdivided into the pleural, pericardial, and peritoneal cavities, enveloping the thorax, heart, and abdomen, respectively.

Then my lecture materials and the research site say

Coelom is the secondary body cavity.

Finnish Wikipedia says that Coelom is only with invertebrates.
Again the Wikipedia page about peritoneum suggests that human has abdominal cavity and no coelom, and other mammalians coelom.

Does human have Coelom?

The confusing thing is the use of the word "OR", since I am not sure whether people are using it in different pages like "XOR" or like "AND" in normal speaking.

molecular biology - How do proteins and genes participate in learning?

The storage of memories in cells is rarely thought of on the protein level of the cell. Cells are usually given a developmental state, but no memory. A cell may become a liver cell, cancerous, or diabetic, but this is not memory, but a physiological change in the cell which is usually not reversible to a previous state.

For example cancer treatments are entirely focused on identifying the cancerous cells and killing them. Internally the genomes of cancer cells often have deletions and duplications. They are cancerous, they have not learned to be cancerous. Though not as dramatic, it is now thought that cellular differentiation which creates different types of cells is heavily influenced by epigenetic modification of the genome; the DNA is marked by methyl groups which dictates the state of the cell by modifying the gene. This is mediated by proteins for sure, but is quite complex and not well understood at this time. Epigenetic markers can even change gene behavior between generations of offspring as well, though that is not usually called memory.

How is information stored in the brain? This is thought to be reflected in the organization of the neurons in the brain. There are many kinds of neurons. They can be distinguished by the sorts of axons and dendrites that emanate from the cell body. They can also be distinguished by the chemical variety of neurotransmitter they use (there are a score of different molecules). So to a great extent the type of cell and the specific proteins it chooses to use to mediate information is very important.

That being said, information is currently thought to highly related to the placement of the axons and dendrites connecting the neuron to sometimes scores of other cells, sometimes touching the cell body, other times other dendrites or neurons. As neural activity ensues, the cells will reconfigure their connections by physically moving them.

More recently, investigators have tried to understand the genes which internally modulate the neural signals within the cells. This nobel prize lecture discusses how the CREB/MAPK pathway can modulate Long Term Potentiation - the shape of the neuron response to a signal over time (days or hours).

Taken as a whole, you can see that memory is likely to be stored on several levels at once - the kinds of cells (dictated by differentiation) involved, structural arrangement of the neurons (axons and dendrites connecting to various cells and places on cells), as well as internal signaling circuitry that generates and modulate the electrical and chemical activity within the cell.

"Marker protein" only refers to a protein that you can follow to see some sort of activity in the cell. A typical example is Green Fluorescent Protein, which is colored, fused with a protein of interest. It has no specific meaning regarding learning I think.