Tag Info

This is particularly an issue when you do maxipreps and the pellet is 200x the size of the pellet from a miniprep. The reasons why? Maybe you're pelleting at too high of a g-force (pure speculation). Alternatively, you could be growing the cells too long. I typically miniprep after a 12 hr culture. If you're seeing any darker colors in your pellet, you have ...

If I recall correctly (from when I interned at ABI), you can't directly set this in the Veriti interface. However, (again, I'm not 100% certain -- I couldn't find anything to confirm this after 30 minutes of googling,) the ramp rate percent refers to the percent of maximum ramp rate for the heating block. According to the product website, the max heating ...

This is a perfectly reasonable result. Remember that you are measuring an average value for a very large population of BSA molecules. Essentially what the 0.4 means is that at any one time, about 40% of the BSA molecules have a free thiol, while the rest of the cysteine residues are disulfide-bonded.

The paper you cite says that the break points are single stranded DNA which have specific proteins bound to them. I'm not an expert here, but if thats the cause of meitotic break points there are some interesting possibilities for detecting them: you could detect them with a tiling array. - that's an micro array which has an oligomer every 40 bp or so ...

As Thomas pointed out, there is a maximum ramp rate which you can set up on the thermocycler. Its value can go from 0% to 100%. I can then assume that a 100% ramp rate will corespond to the maximum ramp rate of 4.25°C/s for the heating block or 5°C/s for the sample. which means that a ramp rate of ~2.4%. Subsequently, I set up my reaction at a 2% ramp rate, ...

Proteinase K activity is greatly increased by addition of denaturing agents like SDS or urea (Hilz et al., 2008), indicating that the denaturation of the substrates helps Proteinase K to degrade them. Increasing the temperature to 50°C will also unfold some proteins already, making it easier for the Proteinase K to degrade them. The proteinase K seems to be ...

We routinely use E. coli preps from 16-18h overnight cultures. The cultures don't show any overt signs of lysis (sometimes you can see cellular debris if the culture has indeed overgrown). I have also noticed inconsistent times when resuspending the pellet. For my own use, centrifuging at too great a g-force or for too long can increase the likelihood of ...

What I do to avoid this is to ressuspend the cells by pipetting up and down the P1 (ressuspension) buffer on the side of the tube. This way, every time takes only a bit of cells every time and big clumps are avoided. The only times I used to get big clumps hard to ressuspend were when I went poking at the pellets first.

The binding of proteins (and cells) to glass (or silicon) surfaces can be prevented by coating the glass with polyethylene glycol (PEG) groups. PEG-silane is a reagent used to create this coating. PEG-silane (the image shows a methoxy- version) (image taken from here; no connection) will coat glass surfaces because the silane portion (right hand end of ...

Look at chemical hydrogen bond breakers. Guanadine hydrochloride is included in the buffer conventionally to compete with the the hydrogen bonds that form the double helix. Other hydrogen bonders like Propionamide and 2-Pyrrolidone can weaken the helix and I would think lower the melting point of the bonds. How much the Tm is affected would depend on the ...

Perhaps isothermal amplification is possible (NASBA)? Amplification of DNA also seems possible (NASBA at biomerieux).

You can look up Gibson Assembly or Circular Polymerase Extension cloning (CPEC). For both of these the website for J5 has some good protocols. Here is the one for CPEC: http://j5.jbei.org/j5manual/pages/80.html For CPEC you can look at the 2011 Quan paper: http://www.ncbi.nlm.nih.gov/pubmed/21293463 Hopefully that helps.

The answer is yes, not only meiotic, but any DNA double-strand breaks will be detected as long as they are processed by homologous recombination. This method (SSDS) is based on the detection of resected ssDNA ends at DSBs. The only difference for the mitotic breaks will be that you cannot use anti-DMC1 for ChIP. Instead, you should use either anti-Rad51 or ...