Prather:DH10B Stocks

Electrocompetent E.coli DH10B cells are used regularly by many members of the Prather Lab. The following rules should be followed when working with competent cells to ensure that these cells are always available in the lab.

How They Arrive From the Supplier

Electrocompetent DH10B Cells (Electromax from Invitrogen) are packaged in 100 ul aliquots, 5 tubes per box.

How They Are Prepared for Use

To use, a tube of electrocompetent cells is thawed on ice, diluted with equal volume (100 ul) cold, sterile 10% glycerol, and distributed in 20 ul aliquots in sterile, chilled 0.2 ml tubes. One tube is used per transformation.

How To Ensure the Stocks Don't Run Out

If you plan to use electrocompetent cells, you should follow these rules:

1. Go to the white box labeled "Electrocompetent Cells" in the -80 freezer. If there are at least N+1 aliquotted tubes, where N is the number you need for a transformation, go for it.
2. If there are N tubes or less, then you have the pleasure of preparing 20 ul aliquots. (That is, don't leave the box empty.) You should remove one (or more) yellow-capped tube(s) from the white box and dilute and distribute as described above.
3. If you remove the last yellow-capped tube from the white box, you should open a new (red) Invitrogen box of cells and transfer all 5 of those tubes to the white box. If there is only one or less new box remaining, add "Electrocompetent DH10B" to the Quartermaster board under "Running Out." This should ensure that we always have sufficient back-up.

A Few Other Notes

• "MaxEfficiency" cells are not electrocompetent, they are chemically competent.
• "MegaX DH10B" cells are not the same as "Electromax DH10B." They use a different recovery medium (not SOC), and might use a slightly different transformation protocol. We did not order these, they were freebies. Feel free to use these if you'd like. The protocol for them is in the drawer with the Qiagen manuals.

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