IGEM:Peking/2007/Switch-Notebook/2007-7-10: Difference between revisions
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Chen Daizhuo (talk | contribs) (New page: Experimenter: LXL, LCB ==Contents== ===Precipitate with ethanol: SDD-EGFP^^, pLX007^^=== * Time: 8:30 ===Redo digestion of pLX007 with XhoI and SalI=== ===Colony PCR test of ECFP-pCC00...) |
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Latest revision as of 07:40, 10 July 2007
Experimenter: LXL, LCB
Contents
Precipitate with ethanol: SDD-EGFP^^, pLX007^^
- Time: 8:30
Redo digestion of pLX007 with XhoI and SalI
Colony PCR test of ECFP-pCC002
- Primers: pkem-F, pCC-test-R
l to r: Marker, ECFP-pCC002(1-15), control
Select for incubation: ECFP-pCC002(6)
图像不清楚大约是因为Bobo错把TAE当了EB+TAE.
Transformation
- SDD’-EGFP-pCC002
- ECFP-pCC002
- pCC002 (C/N)
- CD1-pMD18T
- RD1-pMD18T
Ligation
- MCS-pCC009 4:1
- pCC009 (self ligation control)
Retract of SDD-EGFP^^, pLX007^^
Sequencing result received
Temperature gradient PCR of RD2, RD3
- Template: MG1655 FT Genome
- Prime F: Rec_D_2_F/Rec_D_3_F
- Prime R: Rec_S_2_R (new) / Rec_S_3_R (new)
- Temperature: 50C, 53C, 58C, 63C, 68C
l to r: RD2(1-5), ladder
不浓, RD3也不浓. 问什么我P的就不浓呢? 呼唤NM师兄!
Ligation
- pLX007, lacZa 1:4
- pLX007 (x/s), self ligation control
- pCC002-SDD-EGFP
ExTaq PCR
- 35 cycles
- Product: SS1, SS2, SS3, SD2 (X/S), SD3
- Template: Psul-pMD18T, Psul-pMD18T , Psul-pMD18T, SD2-pMD18T, SD2-pMD18T
- Primer_F: Sul_S_1_F, Sul_S_2_F, Sul_S_3_F, lacO2_XhoI_F, Sul_D_3_F
- Primer_R: Sul_S_1_R, Sul_S_2_R, Sul_S_3_R, lacO2_XhoI_R, Sul_D_3_R
l to r: SS1, SS2, SS3, SD2 (X/S), SD3, ladder
- Retract
- Ligate SS1-3, SD3, pMD18T
要是SD2 XS P不出来就惨了, 我还得重写引物, 明天温度梯度试试吧.
Transformation
- 7 Amp+: SS1-T, SS2-T, SS3-T, SD3-T, CD1-T, RD1-T, pLX007 (xs)自, pLX007-lacZa
- Cm+: SDD_EGFP_pCC002
