Template:Team:UC Berkeley/Notebook/DV construction

K112701
Construction of [] Basic Part K112701

PCR dv001/dv006 on Phns from E. coli K12 MG1655           (~342 bp, gp = A) PCR dv005/dv003 on Phns from E. coli K12 MG1655            (225 bp, gp = B) PCR dv004/dv002 on Phns from E. coli K12 MG1655            (~149 bp, gp = C) --- PCR dv001/dv003 on A+B             (355 bp, gp = D) --- PCR dv001/dv002 on C+D             (697bp, EcoRI/BamHI/DpnI) Sub into pBca1256                  (EcoRI/BamHI, 2472+697) Product is pBca1256-K112701 --- dv001	Forward Biobricking of []     CgATAgaattcATGagatctGAAATATAGCTGTGCCATCAGCC dv002	Reverse Biobricking of []     cagtcggatccGCACGAAGAGTACGGATG dv003 Removing the 2nd EcoRI site in [] (R)  GCCAGGAATGTAAGGgATTCAAAATTGTTC dv004 Removing the 2ND EcoRI site in [] (F)  GAACAATTTTGAATcCCTTACATTCCTGGC dv005 Removing the FIRST EcoRI site in [] (F)CGCTTAATAGGGgATTCTCGTAAACAC dv006 Removing the FIRST EcoRI site in [] (R)GTGTTTACGAGAATcCCCTATTAAGCG

K112703
Construction of His-Tag> basic part K112703

Mix dv009/dv010                       (41bp, EcoRI/BamHI/DpnI) Sub into pBca1256                     (EcoRI/BamHI, 2472+41, L) Product is pBca1256-K112703 --- dv009	Forward Biobricking of His-Tag> cgataGAATTCatgAGATCTatgCATCATCATCATCATCATGGATCCtaacg

dv010  Reverse Biobricking of His-Tag> cgttaGGATCCATGATGATGATGATGATGcatAGATCTcatGAATTCtatgg

K112704
Construction of  basic part K112705

Wobble PCR dv013/dv014               (47 bp, EcoRI/BamHI/DpnI) Sub into pBca1256                     (EcoRI/BamHI, 2472+47, L) Product is pBca1256-K112705 --- dv013	Forward Biobricking of S-Tag> cgataGAATTCatgAGATCTatgaaagaaaccgctgctgctaaattcg dv014  Reverse Biobricking of S-Tag> cgttaGGATCCgctgtccatgtgctggcgttcgaatttagcagcagcgg

K112706
Construction of  basic part K112706

PCR of dv015/dv016 on pBca1037         (500bp, EcoRI/BamHI/DpnI) Sub into pBca1256                      (EcoRI/BamHI, 2472+ 500, L) Product is pBca1256-K112706 --- dv015 Forward biobricking of  cgataGAATTCatgAGATCTccttatgcagcgtaagggccgcaac

dv016 Reverse biobricking of  cgttaGGATCCGATAATGTtTGCAGGGGAATTATTTTG

K112707
Construction of  basic part K112707

PCR of dv017/dv016 on pBca1037        (499bp, EcoRI/BamHI/DpnI) Sub into pBca1256                      (EcoRI/BamHI, 2472+499, L) Product is pBca1256-K112707 --- dv017 Forward biobricking of  cgataGAATTCatgAGATCTaGATCCTGTGATGTTTGGCG

dv016 Reverse biobricking of  cgttaGGATCCGATAATGTtTGCAGGGGAATTATTTTG

K112708
Construction of  basic part K112708

PCR of dv019/dv020 on  MG1655          (225bp, EcoRI/BamHI/DpnI) Sub into pBca1256                       (EcoRI/BamHI, 2472+225, L) Product is pBca1256-K112708 --- dv019 Forward biobricking of  cgataGAATTCatgAGATCTgctaagcgtgaaataccggatgg

dv020 Reverse biobricking of <PfhuA> cgttaGGATCCactctgatgtaaagtgaatgataacg

K112709
Construction of basic part K112709 Wobble PCR of dv021/dv022             (66bp, EcoRI/BamHI/DpnI) Sub into pBca1256                     (EcoRI/BamHI, 2472+66, L) Product is pBca1256-K112709 --- dv021 Forward biobricking of b1006 cgataGAATTCatgAGATCTaaaaaaaaaccccgcccctgacagggcgg

dv022 Reverse biobricking of b1006 GGATCCtaaaaaaaaccccgccctgtcaggggcggggtttttt

K112710
Construction of <Bca1041> basic part K112710

PCR of dv023/dv024 pSB1AG0-Bca1041                   (122bp, EcoRI/BamHI/DpnI) Sub into pBca1256                     (EcoRI/BamHI, 2472+122, L) Product is pBca1256-K112710 --- dv023 Forward biobricking of Bca1041 cgataGAATTCatgAGATCTgCcggcttatcGgtcagtttc

dv024 Reverse biobricking of Bca1041 cgttaGGATCCatttcttttgggtatagcgtcgtgg

K112711
Construction of rbs.SpvR! basic part K112711

PCR of rbs.SpvR! al0027 and al0028  (933bp, EcoRI, BamHI, DpnI) Sub into pBca1256                       (EcoRI/BamHI, 2472+933bp, L) Product is pBca1256-K112711

al0027 Forward Biobricking of rbs.spvR! CCATAGAATTCatgAGATCTAAAAAAGGAGATATTATGGATTTCTTG

al0028 Reverse Biobricking of rbs.spvR! CGttaGGATCCaggtcagaaggtggactg

K112703 again
Construction of His-tag> basic part K112703, 2nd attempt

EIPCR of His-tag> dv025 and ca1246R on pBca1256  (2704bp EcoRI/BamHI,) Cyclize pBca1256-K112703              (EcoRI) Product is pBca1256-K112703

dv025  Forward Biobricking of His-tag>  ctagtGAATTCatgGGATCCtaaCTCGAcgtgcagg ca1246R Reverse EIPCR from mRFP to facilitate Biobricking of His-tag> GACCTTCACCTTCACCTTCG