User:Meng Xiao He/Notebook/fall08/2008/12/10: Difference between revisions

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*use 12-8 protocol on cco brown colonies 1 and 2: each takes cco5F, cco3R, and just pair of 3 outers
*use 12-8 protocol on cco brown colonies 1 and 2: each takes cco5F, cco3R, and just pair of 3 outers
*use Platinum adaptation of 12-8 protocol:
*use Platinum adaptation of 12-8 protocol:
**Cycling conditions:
:Cycling conditions:
#5 min at 94
#5 min at 94
#30s at 94
#30s at 94
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#10 min@ 72 to end
#10 min@ 72 to end
#hold at 4
#hold at 4
==Plating on sucrose==
==Plating on sucrose==
*1:4 dilution into fresh LB, let grow, then plate
*1:4 dilution into fresh LB, let grow, then plate

Revision as of 16:27, 9 December 2008

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PCR tests

  • use 12-8 protocol on cco brown colonies 1 and 2: each takes cco5F, cco3R, and just pair of 3 outers
  • use Platinum adaptation of 12-8 protocol:
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

Plating on sucrose

  • 1:4 dilution into fresh LB, let grow, then plate
  • 40ul ferric citrate added to plates for B colonies
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