Lab June 25 2009

June 25, 2009

We will need to make the following soon (from Dr. Nano):

Make up buffers for the qiagen spin column protocol found on Open wet ware

A liter of each

P1

P2 (store alkalai like this in plastic bottles.)

N3

PE, PB, EB (or 0.1 X TE...TE is used for everything).

LB agar plates; 2 liters worth

500 ml for Ap

500 ml for Km

500 ml for agar with no antibiotic

500 ml for Cm or Tc if needed or Ap.

LB has different formulaes. We'll use compromise: 5 g/l Yeast extract; 5 g/l NaCl; 10 gm/l Tryptone. [edit. Qiagen recommends 10 g/l NaCl, so Nano has said go with that.]

LB broth 2 liters: dispense in 100 ml bottles.

Stocks of Ap, Km, Cm, Tc. Keep in freezer.

Transformation buffer.

Wash/sterilize large centrifuge bottles.

Prepare DEAE paper for agarose gel elution.

Sterilize microfuge tubes; 1.5 and 0.5 ml.

I'll streak DH5 alpha and that strain for ccdB containing plasmids.

Put some of our spin columns in Hcl so you can use them in a day or two.

Someone should take each protocol that we use and make a Word file. Mostly this is just copying the open wet ware and deleting the things we don't use.

and make sure to autoclave lots of wooden sticks (for streaking) and tooth picks (for picking colonies). Make some spreaders too.

and we have chemically competent cells for the ccdB plasmid

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