User:Meng Xiao He/Notebook/fall08/2008/12/10

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PCR tests

 * use 12-8 protocol on cco12.9B, C: each takes cco5F, cco3R, and just pair of 3 outers, 20ul
 * (update: does not work) use Platinum adaptation of 12-8 protocol: 25ul
 * Cycling conditions:


 * 1) 5 min at 94
 * 2) 30s at 94
 * 3) 30s @ 61, 57 for 40 cycles
 * 4) 7min @ 72 (should capture WT cells as well), only 2min for last 12 cycles
 * 5) 10 min@ 72 to end
 * 6) hold at 4
 * 61: cbb12.9A, B, C, cco12.9D; 57: cbb12.9D, E, cco12.9A

on sucrose

 * 1:2 dilution into fresh salt-free-LB, let grow 1hr, then plate 10ul
 * cultures were very thin (may be because had to user rocker- shaker is nonfunctional)
 * cco12.9A grew thick, though

on Gm

 * remaining Gm culture of wash off from filter of Duo


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