1048-pEC-RNaseI
Cloning of 1048-pEC-RNaseI
[edit]Purpose: Cloning of1048-pEC-RNaseI
Date: 2013-3-
Notebook:
Page:
Clone ID:1048
Experimenters: Shanyi
Results
[edit]cloning
[edit]Date:2013-3-
gene synthesis
Fragment1: RnaseI-1 ~RnaseI-20, ~650 bp,
Fragment 2: RnaseI-21~40, ~740 bp,
then use Fragment1+Fragment 2 as template
RnaseI-1+RnaseI-40 as primers
digest :
| band 2,3 |
band 4 | |
| PCR product |
pEC | |
| enzymes |
Nco1+ Xho1 | |
Colony Screening
[edit]Date: 2013
send 1048-1 to sequence, has a G to T mutation and frame shift, need to use use oligo to fix the frame shift and the mutation.
cloning
[edit]pcr
Fragment1: use RnaseI-Nco1-F and RnaseI-18 as primer
Fragment 2: use RnaseI-19 and RnaseI-40 as primer
then use Fragment1+Fragment 2 as template
RnaseI-1+RnaseI-40 as primers
digest :
| band 1 |
band3 | |
| pEC |
PCR product | |
| enzymes |
Nco1+ Xho1 | |
Colony Screening
[edit]Date: 2013
Result intepretation/Summary
[edit]Conclusion
[edit]