SBB10Ntbk-JoseGutierrez
From OpenWetWare
Construction Files
SBB30: 5'UTR_rep009
Biobricking of O99 rep Gene PCR JG001-F and JG002-R on pEC52 (1235 bp, EcoRI/BamHI) Sub into pBca9523-Bca1144 (EcoRI/BamHI, 910+2472, L) Product is pBca9523-sbb30 {5'UTR_repO99!} ---------------------------- JG001-F Cloning of O99 rep Gene ccataGAATTCatgAGATCTaaaaacgattctgacgcattttttatg JG002-R Cloning of O99 rep Gene CTGATGGATCCCTACTCTACAAGACCTCGTTTTttc
SBB38: PBad Promoter
Eco/Bam transfer pBjh1601CK-ig114 Sub into pBjk2741-Bca1144 (EcoRI/BamHI, 910+2170, L) Product is pBjk2741-ig114 {AraC-Pbad}
SBB37: PCon Promoter
Eco/Bam transfer pBca1100-Bca1152 Sub into pBjk2741-Bca1144 (EcoRI/BamHI, 910+2170, L) Product is pBjk2741-Bca1152 {Pcon-J23100}
Parts Assembly
2/17/10
Objective: Set up PCR reaction for biobricking of SBB30 009 rep Gene -PCR JG 001-F & JG002-R on pEC52 (1235 bp Eco/Bam)
Procedure:
- convert Oligos to 100uM [X(nmol) + 10*X(ul)]
- dilute stock Oligos to 10uM
- PCR Procedure
2/17/10
Objective: Run Analytical gel on SBB30 (JG01), after confirmation run zymo cleanup
Results:
Dorothy
- ladder
- 28
- 29
- sbb25
- sbb26
- sbb32
- sbb34
- JG1
- ZHH 1,2 AG
- TN 001/002 PT
- JHA
- JHB
- JHC
PCR product confirmed, continue to Zymo Cleanup
2/24/10
Eco/Bam Digest of SBB30 PCR products
3/1/10
To Do List: Zymo gell purification of Digest
Eco/Bam transfer pBjh1601CK-ig114
Sub into pBjk2741-Bca1144 (EcoRI/BamHI, 910+2170, L)
Product is pBjk2741-ig114 {AraC-Pbad}
Eco/Bam transfer pBca1100-Bca1152
Sub into pBjk2741-Bca1144 (EcoRI/BamHI, 910+2170, L)
Product is pBjk2741-Bca1152 {Pcon-J23100}