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.