CharonY

Rare relative to what?

It depends quite a bit on the protocol (and kit components). I always made the stains myself and use the MS-compatible staind. One of the first things to go tends to be the sensitizer. If made yourself or the bottle has been opened (but stored appropriatelyI would think that within a few months the components would degrade significantly. Fully closed kits tend to have shelf lives of 1-2 years (though I would check the manual and lot information). 3-4 years may be pushing it (especially if they have been around a while before purchase), but I would simply run a test gel with a dilution series of a reference sample and look how sensitive it still is and whether it is sufficient for applications.

It is also quite a bit dependent on perspective. Scientists from different (sub-)discipline. From a physiological perspective the lack of own metabolic activities is probably the most important distinction (whereas symbionts engage in metabolite exchange, for instance).

Excellent point!

Well, GCMS is more for identifying metabolites, as many of the more interesting compounds tend to be volatile (terpenes, flavenoids and lipids come to mind). But isolating them would be difficult endeavor with that, I agree. It really depends on what you want to do, but for a metabolomic approach I would do an extraction and run it on a GCMS to see whether there is anything of interest to begin with (which can be coupled with pre-fractionation). Especially because a decent GCMS is generally way cheaper than a comparable LCMS.

If you need to identify compounds in complex mixtures I do not think there is any better way than LCMS or GCMS. Preparative HPLC can give you very good fractionation, provided that the LC part is set up very accurately and the complexity of your mixture does not overwhelm its separation and fractionation capabilities. Unless there is further purification and prefractionation a complete plant extract would be highly complex, however.

With regards to autoclaving, you can do that. Just be aware that oftentimes autoclaves are not very clean and it may contaminate your solution (although it would be sterile contamination). Depends on the sensitivity of your application. If made cleanly Tris tends to be fairly stable at RT (6 months did not appear to be much of an issue), though I would check pH before use to be certain. Also note that the pH is temperature dependent. Should you put it in the fridge, let it warm up to RT (or whatever the reaction temp is) before use.

Welcome to the wonderful and expensive world of molecular biology. The moment you deal with complex mixtures, the cost skyrocket. With low complexity and well-known compounds you may get away using chromatographic methods with simple detectors.

Now that got incoherent pretty quickly. But in case you wanted to know, bacteria actually produce D-amino acids (e.g. as elements during peptidoglycan synthesis). And as I said, there are enzymes responsible for the interconversion of the amino acid from the D to the L form.

Both amino acids are found in nature, but only L-amino acids are found in proteins. Biosynthesis amino acid can lead to both forms. Transaminase are pretty much unrelated to this question, as they catalyze the exchange between amino and a keto group. Think isomerase.

As BabcockHall mentioned, the regulation is enacted via proteins, not directly via the steroids/hormones (most notably proteins of the nuclear receptor superfamiliy). There are therefore multiple layers that modulate the final expression. One is on the level of the protein with all the possibilities such as by abundance, inhibition, activation, etc. On the metabolite level inactivation of thyroid hormones (I assume we are talking T3/T4 here) there are several pathways but generally is a form of deiodination.

A bit of a tangent, but it isn't it weird to propose to ban information on things that are relatively easy to design even without a detailed manual , whereas purpose-made devices for causing harm (such as guns and ammunition) are freely available? Isn't that a bit putting the cart in front of the horse?

Whether a fragment is sticky or blunt basically depends on how the respective strands are nicked. In most cases star activity changes the sites only by a little (they still tend to be palindromic to some extent, for example) chances are that they will produce same ends as they did under normal circumstances. Due to the loss of specificity the overhang sequence may very well be different, however.

As I mentioned, I do not think that anyone has investigated the precise physics behind it in detail. The best I found was more or less a list of potential interactions with all the usual candidates (with hydrophobic interactions being near the top of the list). There are some models around, mostly from the manufacturers as e.g.: http://www.pall.com/main/oem-materials-and-devices/literature-library-details.page?id=27294.

Just wondering, how is that different from standard optical tweezers (yeah I know, I should read the articles...)?

I think the question is actually: "what is consciousness"? It may very well be an emergent property of how our brain works and does not need to be beneficial per se. Also consciousness is a physical process (as evidenced by by lesion studies). So why the distinction?

RNA is mostly present as single-strand, but it can form secondary structures with itself or other nucleic acids. Thus a certain level of denaturation is required to prevent that. The role for SDS in proteins serves several purposes, actually, of which denaturation is only one.

Great, now you made my eyes bleed.

Well, typical cyanobacteria have doubling times of a few hours...

Also smells like homework.

For simplicity in terms of cultivation I would probably go for cyanobacteria. Some have engineered growth vessels which maximize surface so that the little buggers can get sunlight very efficiently. Nonetheless, the yield for use as biofuels is still very low, even with them. It may depend very strongly on how efficiency is measure, but the conversion efficiency of sunlight to biomass via photosynthesis is closer to 2 (and lower) %, from what I have seen.

1) If you look at database entry you will generally see the complete ORF. Depending on what you need (e.g. if you also want up- and downstream sequences) you may have to switch viewers that provide genomic views and you may need to search a little bit (using the annotated genome sequence). 2) There a hundreds of websites discussing primer design. Much can depend on the type of fragment you want to amplify (e.g. length, extensions, tags etc) as well as the polymerase. But for a general primer on that topic I would just google it (e.g. http://www.premierbiosoft.com/tech_notes/PCR_Primer_Design.html) and see what applies to you.

it is usually a non-issue as the efficiency of the whole cloning experiments tends to be way lower than that of the mismatch rate. You have a slightly higher chance of creating some false products, but depending on how the designers are designed and how the cloning is going to proceed you will end up with the vast majority of the correct product for cloning. Since you have to run controls anyway, the probability of PCR purity being the culprit for failed cloning is very low. There are exceptions however, e.g. if you do some tricky amplifications with very low yields. Salts or impurities themselves are no issue at all as you do a cleanup post-PCR, anyway.

I do not think that this can be easily answered. The strength of the immune responses is determined by a multitude of factors, for example. Likewise normally the respective antibody levels will decline and can fall below detectable levels. Though it does not mean that there are no cells left that present them. In addition, antibody reactions are no necessarily 100% specific, meaning that very weak responses may be triggered with structurally similar compounds. In cases of strong reactions (e..g due to certain infections) especially IgMs can be readily detected after a year or so. More sensitive methods may even spot them longer.

Well, there are labs specialized in monitoring sleep. But with insomnia it is important to understand whether you are truly fully awake or just have a bad quality of sleep, during which you just feel awake. Sleep monitors can assist in figuring that out (or, even better, look for a specialist). There are a lot of guides and help out there to improve sleep hygiene, but it depends a lot on the reasons for the bad quality of sleep/insomnia to begin with. It could be worthwhile to checking those out, though (as following them is certainly not harmful).