CharonY

If such a high overhang is really needed first check if it makes stemloops. What is the calculated annealing temperature for the primers? What you could also try in theses cases are typically betaine and DMSO.

Basically that is correct. In about any case I can think of the virus attaches to certain receptors. The mode in which the virus inserts genetic material varies though, as described above.

It also depends on the organism/tissue you want to isolate the DNA. For PCRs the above cooking lysis is often sufficient, for blots or other things higher purity is needed. Most protocols not using columns incorporate a phenol step to remove proteins and similar stuff.

Dak, journals typically only publish the results. Although in some cases I have heard that they also pay other labs for verfication of certain results (only heard it from a physics journal, though. not really my field of expertise). In almost all cases (at least of journals I know of) the editors are scientists themselves, though. Of course they are responsible for the review process, however the reviewers are not paid. More recently some online journals indeed started to appear. Most of them allow free access to the articles, but still charge money from the authors (often less, then many print journals though). I do have the feeling that it is not always about covering costs from the journals as the charges can vary extremely.

Well, the problem is simply that your definition does not match the actual termini used to describe evolution. And they were in fact misleading. There are for instance bacteria that have adapted to parasitic life-styles and in that process eliminated a lot of its own core genome (a more dramatic examples are probabl plastids and mitochondria). In your definition this would be accounted as "backward" or at least as de-evolution. However the process is clearly evolution and not the reversal of it. As such your definitions are simply not suited or accurate enough to describe this. Sorry.

I am still a postdoc (which sucks btw) working in the field of, well, -omics. I.e. genomics and postgenomics (including transcriptomics, proteomics and a wee bit of metabolomics). Almost exclusively on bacteria. While it is fun to work with them, funding is a little bit tricky. Impact factors for instance are much lower for microbiology papers than, say, cell biology ones.

And in addition to page limits there are page charges. I think for my last paper we had to cough up over 2k euros. I should learn to use fewer, or at least shorter words. Ow and by reading swansont's post I have to add, take breaks every few hours. Your wrists and back will love you for this

Ah, OK cross-faculty theses are little bit harder, I suppose. However, most reputations of mean examiners tend to be exaggerated (well, I managed to get some diploma students crying myself). But if he is a real meanie, well you are the expert in your field. He is not

Actually it is traditional to write up till the last minute. On the other hand, I found writing my phd thesis (Ok, it is a little while back, but not that long that it is totally blurry), somewhat less tedious than writing the papers. I mean the thesis only goes to your prof and other usually benevolent reviewers, while the latte actually tend to go to experts and more often than not competition in the same field ;-P

I just wanted to point out that this is not evolution per se. What you describe there is mutation plus natural selection, a mechanism of evolution. Evolution cannot be ascribed to single entities (what you said with "thos who have evolved...") but only to complete populations. As has been said before, evolution is the change of allele frequency in a population. Therefore the occurance of mutations (be it beneficial or otherwise) does not equal evolution. Only if these new alleles spread through the population do we observe evolution. As such there cannot be individuals in a population that have evolved and other that have not.

Precisely. I suppose this misconception is partly derived from the image of evolutionary trees. They only depict the history of species and not a trend of evolution towards anything. Furthermore, natural selection alone does not likely lead to speciation. Spatial separation for instance, maybe together with genetic drift (see allopatric speciation) are far more likely to contribute to this. If for instance the whole population adapts to a given environment there won't be a speciation event as per definitionem no new species arises.

Drats, I missed doomsday.

I suppose you are refering to WWII. As far as I know the belief that the Japanese were some kind of chosen or descendents of some divinity were used to justify slaughters on "lesser races", like e.g. the Chinese. But actually I cannot find any current polls either.

It does not look like a pure culture, at least.

Well, I dunno, I find cultivating bacteria extremely boring. Especially if you have to deal with slow-growing anaerobes. And that is just to get to the time-consuming boring actual experiments. Having the data on the hand...hmmmmmm:D To be a scientist, I think, you have to have a drive for it in the first place. I mean the pay is bad, the workload is high and as in most other professions a lot of it is probably boring routine work. Then it is fairly easy to overcome the boring parts (especially if the alternative is having no job that is )

That's very unusual. Such prolonged sleep deprivation should lead to severe health problems. I get around 4h sleep a day (do to a lousy stess-full job) for something more than two years now. And apparently getting some symptoms already (including migraine, slight cardiovascular problems etc.). I should learn to sleep again. On topic: the longest time frame completely without sleep was around 71-76 hours (thrice) due to a lab-experiment that required constant supervision.

There are gazillions of DNA isolation protocols, depending on the organism (sometimes even lab), though most basics are the same. I could answer more specific if the complete protocol is given. Do you use TES to resuspend your cells? In that case it could be use to prevent osmolysis. Alternatively there are certain columns to which DNA binds with a high salt-content buffer for instance...

Spores don't propagate and can't be thermophilic per definitionem. They are thermo tolerant, however Regarding thermophilic pathogens, thermophilic Campylobacter species comes to my mind there are also some clostridia that have been classified as thermophilic.

Actually I think I read somewhere that annealing of the DNA-templates (that is, forming of hydrogen bonds) is a (the?) rate limiting step in this reaction. At high temperatures these bonds are naturally less stable or easier disrupted. Of course, a host of protocols exist and depending on your templates it might not be essential at all. With sufficient vector and insert e.g. an 1-2 h incubation on the bench usually works. On the other hand I often do o/n incubations in thermos flasks in the fridge for blunt end ligation. Using the thermos flask the temperature decreases more slowly and supposedly increases efficiency...

How about checking with a microbiologist at the nearest university? Setup is pretty straightforward. A standard test is to plate the bactaeria on an agar plate and put discs with antibiotics on it. Measure the zone of inhibition. Each discs contains a different AB (or concentration). Another standard test are MIC-tests (Minimum inhibition concentration). THese are more work. You need liquid medium, each containing rising concentrations of AB. You measure the optical density (OD) to determine which cocentration of a respective AB inhibits growth. Note that in many countries for safety reasons you are required to kill the bacteria after use, which requires autoclaving.

Well there is actually quite a a bit research regarding introns and their function and it is now more or less known that they do carry information that will not directly transform to proteins (that is, get translated). And it ain't some central "dogma". The only dogma i am aware of is the information flow from DNA to mRNA to protein (although in case of reverse transcrpitase activity the information of mRNA might actually lead back to DNA). Same goes to so-called "junk" DNA. For quite a few of them ideas begin to develop what they might be fore (and as such the term junk is clearly a misnomer). Especially the research on small RNAs has led to the discovery of new regualtory roles (or in some cases, splicing functions). Check this for instance http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=14550267

One can either add the substances to the (slightly cooled) agar while before pouring it into the petri dish). Alternatively there are antibiotic discs to test them. Or you can put a sterile filter disk on a plate with bacteria and drop around 10µl on it. For E. colis you might want to ask nearby universities, however you won't (or at least shouldn't) get Streptococci or Salmonella. Both species are usually human pathogenes and may not be cultivated outside specially equipped laboratories! Edit: forgot to add: most chemicals can be acquired by e.g.Sigma, but you may need an authorization to buy there (depends a bit on local laws).

I suppose you mean a change in protein abundancy? That's theoretically possible but for a true quantification you got to use techniques like radioactive labelling or DIGE, or the usage of heavy nitrogen. With 2D PAGES as compared to 1D you get a better resolution and therefore you can visulize more proteins. And in fact there are plenty of papers regarding the proteome of fungi. I don't think that there are 2d gels only woth ribosomal proteins around though, since one of the strength of this technique is the visualisation of a lot of proteins in one gel. If one could separate the ribosmal proteins from all others (at the moment i'd have no clue how short of His-tagging and purifying them) an 1D PAGE should suffice.

None of these qualify as new species. They are just mutant strains.