SBB09Ntbk-Sadao Ota: Difference between revisions
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==[[User:Sadao Ota|Sadao Ota]] 15:03, 25 February 2009 (EST)== | ==[[User:Sadao Ota|Sadao Ota]] 15:03, 25 February 2009 (EST)== | ||
'''[[SBB09_Ligation of EcoRI/BamHI digests | Ligation of EcoRI/BamHI digests]]''' | |||
I used gel to separate the DNA I need | I used gel to separate the DNA I need |
Revision as of 13:11, 25 February 2009
Sadao Ota 15:03, 9 February 2009 (EST)
My project is SBB09_20154, to make ag43 Display (short) ag43 Display (long) systems. Designed a oligo M10034 to make Autotransporter protein with promotors and so on (corresponding to amino acids 708-1017). Also, designed another oligo M10035 to make Autotransporter protein only (corresponding to amino acids 517-1017).
- Construction files are M10034 M10035
- Oligonucleotide sequences are Oso001, Oso002, Oso003
Sadao Ota 15:03, 16 February 2009 (EST)
Do PCR with the combination of Oso001&Oso002, and Oso002&Oso003.
Sadao Ota 15:03, 23 February 2009 (EST)
Clean up PCR products for gelation analysis
Clean up PCR products, Digest, Clean up and gel again.
Regular Zymo Cleanup (removes the polymerase, dNTPs, buffer, and most of the oligonucleotides from a PCR reaction) here
Load to Gel to perform gelation analysis (dead or alive) here
EcoRI/BamHI Digest of PCR Products (my PCR products are large enough, so no need to modificate the process) here
Sadao Ota 15:03, 25 February 2009 (EST)
Ligation of EcoRI/BamHI digests
I used gel to separate the DNA I need
I also used digested DNA without gelation to do heat shock.
2nd PCR cloning did not work. So, PCRing again...