Template:Team:UC Berkeley/Notebook/SC construction

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pBca1256-K112600

Construction of <AP-tag> basic part K112600

Wobble PCR SC001/SC002  (66 bp, EcoRI/BamHI)
Sub into pBca1256           (EcoRI/BamHI, 2227+ 910)
Product is pBca1256-K112600
----------------------------------------------------
SC001F    Forward Biobricking of <AP-tag>
cgataGAATTCatgAGATCTggcctgaacgatatttttgaagcgcag
SC002R    Reverse Biobricking of <AP-tag>
ccagtGGATCCttcatgccattcaattttctgcgcttcaaaaatatcg

pBca1256-K112601

Construction of <AP-tag! basic part K112601

Wobble PCR SC001/SC003 (69 bp, EcoRI/BamHI)
Sub into pBca1256           (EcoRI/BamHI, 2227+ 910)
Product is pBca1256-K112601
--------------------------------------------------
SC001     Forward Biobricking of <AP-tag!
cgataGAATTCatgAGATCTggcctgaacgatatttttgaagcgcag
SC003     Reverse Biobricking of <AP-tag!
ccagtGGATCCttattcatgccattcaattttctgcgcttcaaaaatatcg

pBca1256-K112602

Construction of <FLAG-tag> basic part K112602

Wobble PCR SC004/SC005 (45 bp, EcoRI/BamHI)
Sub into pBca1256           (EcoRI/BamHI, 2227+ 910)
Product is pBca1256-K112602
-------------------------------------------------------------
SC004     Forward Biobricking of <FLAG-tag>
cgataGAATTCatgAGATCTgactacaaggatgacgacg
SC005     Reverse Biobricking of <FLAG-tag>
ccagtGGATCCcttgtcgtcgtcatccttgtagtc

pBca1256-K112603

Construction of <FLAG-tag! basic part K112603

Wobble PCR SC004/SC006 (48 bp, EcoRI/BamHI)
Sub into pBca1256           (EcoRI/BamHI, 2227+ 910)
Product is pBca1256-K112603
-------------------------------------------------------------
SC004   Forward Biobricking of <FLAG-tag!
cgataGAATTCatgAGATCTgactacaaggatgacgacg
SC006   Reverse Biobricking of <FLAG-tag!
ccagtGGATCCttacttgtcgtcgtcatccttgtagtc

pBca1256-K112604

Construction of {rbs_barstar>} basic part K112604
PCR mea035/sc007 on Bacillus amyloliquefaciens gen.           (320 bp, EcoRI/BamHI)
Sub into pBca1256                                             (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112604
-------------------------------
mea035 Fwd biobricking of {rbs_barstar>} 
cgataGAATTCatgAGATCTcataagaaaggagccgcacatg                              
sc007 Reverse biobricking of {rbs_barstar>}
cgttaGGATCCagaaagtatgatggtgatgtcgc  

pBca1256-K112403

Construction of {prepro>} of PhoA basic part K112403          (114 bp, EcoRI/BamHI)
CR cb10007/cb10008 on MG1655                                   (EcoRI/BamHI, 2472+910, L)
Sub into pBca1256
Product is pBca1256-K112403                                            
-------------------------------------------------------------
cb10007-phoA-F  Fwd biobricking of {prepro>}
cgataGAATTCatgAGATCTgtacatggagaaaataaaatgaaacaaagc
cb10008-phoA-R  Reverse biobricking of {prepro>}
cgtagGGATCCgggcttttgtcacaggggtaaac


pBca1256-K112404

Construction of {prepro>} of LamB basic part K112404              (132 bp, EcoRI/BamHI)
PCR cb10009/cb10010 on MG1655                                     (EcoRI/BamHI, 2472+910, L)
Sub into pBca1256                                                               
Product is pBca1256-K112404                                           
-------------------------------------------------------------
cb10009-lamB-F Fwd biobricking of {prepro>}
cgataGAATTCatgAGATCTgaaaagcaatgactcaggagatag
cb10010-lamB-R   Reverse biobricking of {prepro>}
cgtagGGATCCcagccattgcctgagcagacattac


pBca1256-K112605

Construction of {a~prepro>} of PhoA basic part K112605             (100 bp, EcoRI/BamHI)
PCR sc008/cb10008-phoA-R                                           (EcoRI/BamHI, 2472+910, L)
Sub into pBca1256
Product is pBca1256-K112605
------------------------------------------------------
sc008 Fwd biobricking of {a~prepro>}
cgataGAATTCatgAGATCTtaaaAtgaaacaaagcactattgcac
cb10008-phoA-R Reverse biobricking of {a~prepro>}
cgtagGGATCCgggcttttgtcacaggggtaaac

pBca1256-K112606

Construction of {a~prepro>} of LamB basic part K112606                 (111 bp, EcoRI/BamHI)
PCR 009/cb10010-lamB-R                                                 (EcoRI/BamHI, 2472+910, L)
Sub into pBca1256
Product is pBca1256- K112606
--------------------------------------------------
sc009 Fwd biobricking of {a~prepro>}
cgataGAATTCatgAGATCTtagaatgatgattactctgc
cb10010-lamB-R Reverse biobricking of {a~prepro>}
cgtagGGATCCcagccattgcctgagcagacattac

pBca1256-K112607

Construction of {rbs_prepro>} of PhoA basic part K112607               (113 bp, EcoRI/BamHI)
PCR sc010/cb10008-PhoA-R                                               (EcoRI/BamHI, 2472+910, L)
Sub into pBca1256
Product is pBca1256-K112607
------------------------------------------------------
sc010 Fwd biobricking of {rbs_prepro>}
cgataGAATTCatgAGATCTgtacatggagaaaataaaAtgaaacaaag
cb10008-phoA-R  Reverse biobricking of {rbs_prepro>}
cgtagGGATCCgggcttttgtcacaggggtaaac

pBca1256-K112608

Construction of {rbs_prepro>} of LamB basic part  K112608         (126 bp, EcoRI/BamHI)
PCR sc011/cb10010-lamB-R on E.coli K12 MG1655                 (EcoRI/BamHI, 2472+910, L)
Sub into pBjh1601AK
Product is pBca1256-K112608
------------------------------------------------------
sc011 Fwd biobricking of {rbs_prepro>}
cgataGAATTCatgAGATCTcaatgactcaggagatagaatgatgattac has little longer annealing region. If you use shorter, GC content goes up little. I would use this:

cgataGAATTCatgAGATCTcaatgactcaggagatagaatg

cb10010-lamB-R Reverse biobricking of {rbs_prepro>}
cgtagGGATCCcagccattgcctgagcagacattac



Sherine Cheung
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