IGEM:Peking/2007/Switch-Notebook/2007-7-6

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Experimenter: YT, YL, LCB, WMC

Contents

Contents

Precipate pLX007(SalI/XhoI), pCC009(SalI/XhoI) and lacZa(SalI/XhoI) with ethanol

2倍体积乙醇, 1/10体积NaAc.

回收于20uL, 离心30min.

Positive transformation of SDA-EGFP-pCC002(1,5) grown up

但提质粒用的是存下来的菌液, BamHI/XhoI(7uL/20uL)切检验, OK! 存质粒.

Image:PKU Switch 07-7-6 SDA-EGFP-pCC002(1,5).gif


l to r: 1, 5, none, Marker


还有菌液在冰箱门上存着, 用时直接提.

mRFP-PCC002, SDY-ECFP-pCC002 plates do not grow, 昨天晚上连的体系转化

RD1-1-T, RD1-2-T transformation

Extract <bbpart>BBa_I13522</bbpart>(GFP)(1-4) from iGEM DNA Kit

Waiting for Mini-prep in the evening, labeled GFP-1,2,3,4.

Done.

Single digestion of pCC009(1-4), pLX007(4) with XhoI (2uL/20uL) to check

lacZa(SalI/XhoI)电泳看浓度, 10uL, Marker 10uL (稍大)

连接

  • lacZa-pLX007(4uL/1uL), H2O-pLX007自连

Image:PKU_Switch_07-7-6_pLX007(4)pCC009(1-4)XhoI.jpg

l to r: pLX007^, pCC009(1-4), pLX007, Marker

  • MCS-pCC009(4uL/1uL), H20-pCC009自连

Image:PKU Switch 07-7-6 lacZa xs.gif

l to r: lacZa(x/s), Marker

5pm-6pm: 4 degree, 6pm-9pm 16 degree.

KOD PCR SD1&2&3

结果如下, 切胶回收SD1&2较大片段, 30uL洗脱, Marker 10uL

Image:PKU Switch 07-7-6 SD1&2&3.gif

l to r: SD1, SD2, SD3, M

连pMD18T (1hour, 16 degree), 转化

Miniprep of <bbpart>BBa_I13522</bbpart>(GFP)

OK! Stock in -20C fridge.

30uL/2uL电泳看浓度

Image:PKU Switch 07-7-6 I13522-gfp(1-4)SD1&2.gif

l to r: I13522(GFP)(1-4), SD1, SD 2, Marker

菌P(分区PCR)

  • RD1-1-pMD18T 1-12区
  • RD1-2-pMD18T 1'-12'区

(每区3-5个不定, 以戳的洞为准)

Looking forward

检板子

  • mRFP-pCC002
  • SDY-ECFP-pCC002
  • SD1-pMD18T
  • SD2-pMD18T
  • MCS-pCC009
  • lacZa-pLX007

菌P结果电泳

  • RD1-1-pMD18T
  • RD1-2-pMD18T

如有对的提质粒酶剪切

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