1383 pCMV5-TPA-NPR1-ECD 2-Fc(3c)-WPRE-OriP
Cloning of 1383-pCMV5-TPA-NPR1-ECD#2-Fc(3c)-WPRE-OriP
[edit]Purpose: Cloning of 1383-pCMV5-TPA-NPR1-ECD#2-Fc(3c)-WPRE-OriP
Date: 2012-9-29 ~
Notebook:0
Page:
Clone ID: 1383
Experimenters: Paul & Shanyi & Xiuli
Results
[edit]PCR
[edit]2012-9-29
10-01-2012-Xiuli
PCR again, 1382 1383 didn't work.
2012/9/30 digestion:
| Lane |
1 |
2 |
3 |
4 |
| Name |
TagGFP |
1369 |
1376 |
TagGFP |
| Digestion |
Nhe1+Not1 |
Nhe1+Not1 |
Nhe1 + Kpn1 |
Nhe1 + Kpn1 |
Colony Screening
[edit]Date: 10/7/2012
should be 1321 bp, is clone $1 correct size?
10-8-2012
Most Miniprep is vector itself, re-screening colony with PCR methods.
No right PCR band shown for 1383.
10-8-2012
Re-do PCR using Kinase 7901 (B07) as template (use miniprep as template before). No. 2 PCR reaction including 2 ul DMSO works and 25 Cycle was used for PCR.
Digest half of gel purified PCR products (10/20ul) and vector 1391 with NheI and NotI,
make ligation with half 1382(1383) digestion and half 1391 vector piece.
10-10-2012 miniprep
Enzyme digestion: NheI+NotI , release 1.3kb band.
All clony looks positive.
Result intepretation/Summary
[edit]