Tag Info

I believe Addgene send strains as bacterial stabs. This is the protocol from Qiagen. E. coli strains can also be stored for up to 1 year as stabs in soft agar. Stab cultures are used to transport or send bacterial strains to other labs. Prepare stab cultures as follows: Prepare and autoclave LB agar (standard LB medium containing 0.7% agar) ...

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 ...

E. coli is mostly harmless; only a few strains are harmful. I don't believe the route by which gut biota is established has been entirely established for any species but, for example, koala feed their faeces to their offspring to help them establish biota capable of digesting eucalyptus. It seems that a small proportion of ingested bacteria somehow survive ...

The cited article describes effects of Tris on the outer membrane of E. coli. Like all Gram negatives E. coli has an inner (or cytoplasmic) membrane which is a typical phospholipid bilayer membrane containing many membrane proteins including transporters and, of course, an electron transport chain. Surrounding this there is a layer of peptidoglycan, and then ...

Not sure that I have ever come across trace metals in recipes (I assume that you aren't referring to the Ca/Mg salts component of M9), but it is certainly the case that many K12 strains (e.g. C600, DH5α) require thiamine because they are thi mutants. This doesn't apply to BL21 since it is from a different lineage (it is a B strain). In a lab where ...

I would rinse the cells off the agar using a small volume of LB or similar rich medium. Then use a small aliquot (10 μl) to inoculate a 5 ml culture under selection to see if anything will grow. Meanwhile the remainder of the cell suspension from the vial can be used as input for your favourite miniprep procedure. If necessary, the DNA that you recover ...

Over time I suspect that ice crystals will form in the colonies from defrost cycles in the freezer and the moisture in the agar. This will cause rupture of the bacteria membranes and over time may destroy the bulk of the bacteria cells. The plates will also desiccate over time as well from the defrosting cycles of the freezer. You may be able to leave ...

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.

Trehalose[1],[2] is the cryoprotectant used by bacteria to keep things going when all else freezes (or dries up), and they grow in it, too! Trehalose accumulates dramatically in microorganisms during heat shock and osmotic stress and helps protect cells against thermal injury and oxygen radicals. Here we demonstrate an important role of this sugar ...

I think of E. coli being more white than yellow. When you compare E. coli and S. aureus on an agar plate, the Staph colonies have a much more distinctive yellow tint. The yellow tint in Staph colonies is due to staphyloxanthin, a carotenoid pigment.