IGEM:Peking/2007/Switch-Notebook/2007-7-8

Experimenter: LXL, NM, CCY

pMDB与pMDB0的效率差别不大
(当然还是少了一点的)

看来感受态放在-40C还是可以的, 上次应该是运输的问题, 下次可以用liquid nitrogen直接带来

Colony PCR test of SD1,2, lacZa-pMD18T

 * primers: M13F/R
 * control: positive transformation of pMDB



l to r: Marker, Negative control, SD1(1-12), Negative control, SD2(1-10)



l to r: Marker, SD2(11-12), Negative control, lacZa(1-11)

For sequencing tomorrow: SD1(1,4,5), SD2(1,2,3)

For miniprep tonight: lacZa(1,2,3)

ECFP^^失活, precipitate with enthanol

 * precipitate at 9:30am
 * collect at 2pm

Miniprep of ECFP1-5, mRFP1-20, 酶切检测


l to r: ECFP(1-5), mRFP(1-19), Marker


 * ECFP is not ligated.
 * mRFP(2,4,14,18) are correct.

SDE-EGFP-pMD18T酶切


l to r: Marker, mRFP20, SDE-EGFP-pMD18T^^

What’s wrong?

Re-try:



l to r: Marker, SDE-EGFP-pMD18T, ECFP^^

OK!

KOD PCR of SDD, SDD’_EGFP

 * Primers: SDD_EGFP_BamHI_F, SDD’_EGFP_BamHI_F (newly ordered) + GFP-ClaI-R
 * Template: SDD_EGFP_pMD18T



l to r: SDD, SDD', none, Marker, SDE-EGFP-pMD18T^^

切胶回收, 双切过夜 (BamHI & Cla I)

Result of pCC002-ECFP
Failed. pCC002自连太多, 于是失败. pCC002重切过夜.

ECFP^^回收, 连接, 过夜

 * ECFP, pCC002, ClaI/NotI

Miniprep of pLX007, lacZa (xs)-pMD18T
No SalI.