From OpenWetWare
Construction of FokI Cleavage Domain basic part sbb19
PCR mb0001/mb0003 on BBa_K157000 (504 bp, gp = A)
PCR mb0002/mb0004 on BBa_K157000 (143 bp, gp = B)
---------------------------------------------------
PCR mb0001/mb0004 on A+B (622 bp, EcoRI/BamHI)
Digest pBcajk2741-Bca1144 (EcoRI/BamHI, 2170+910, L)
Product is pBjk2741-sbb19 {<FokI+!}
---------------------------------------------------
mb0001 Forward EcoRI + BglII
ctacgGAATTCgctAGATCTCAGCTGGTTaaatctgaactggaggag
mb0002 Forward point mutation A-->T cataaaaccaatTgcaatggcgccg
mb0003 Reverse point mutation A-->T cggcgccattgcAattggttttatg
mb0004 Reverse BamHI gtacgGGATCCTTAaaaattgatctccccattg
JCA Notes
- Fixed wiki link
- Prefix should be ATG not GCT
- Changed template name to partName
- Construction file is correct otherwise
- Changing some oligos for sharing with another part
Construction of FokI Cleavage Domain basic part sbb19
PCR fk0001/fk0003 on BBa_K243000 (504 bp, gp = A)
PCR fk0004/mb0002 on BBa_K243000 (144 bp, gp = B)
---------------------------------------------------
PCR fk0001/mb0002 on A+B (622 bp, EcoRI/BamHI)
Digest pBjk2741-Bca1144 (EcoRI/BamHI, 2170+910, L)
Product is pBjk2741-sbb19 {<FokI+!}
---------------------------------------------------
fk0001 Forward EcoRI and BglII for FokI
ccaaaGAATTCatgAGATCTCAGCTGGTTaaatctgaactggaggag
fk0004 (F)Base Substitution FokI
ccataaaaccaatTgcaatggcgccg
fk0003 (R)Base Substitution FokI
cggcgccattgcAattggttttatgg
mb0002 Reverse BamHI
gtacgGGATCCTTAaaaattgatctccccattg
