2321 pFastBacDual-VSVG-CAG-Flp
Cloning of 2321 pFastBacDual-VSVG-CAG-Flp
[edit]Purpose: Cloning of2321 pFastBacDual-VSVG-CAG-Flp-==
Date: 2014-9----
Notebook:0
Page:
Clone ID:2321
Experimenters: shanyi
Results
[edit]CLONING
[edit]Use 503 psPAX2 as template, PCR CAG promoter
2321-CAG-Nhe1-R (ctgtctcatcattttggcaaa gaattc ggatcc gcc acc atg gct agc tagca)
tgctagctagccatggtggcggatccgaattctttgccaaaatgatgagacag
2321-CAG-BglII-F tactaAGATCTgtcgacattgattattgactag
Digest with BglII + Nhe1.
Digest vector with BamH1 + Nhe1.
Colony Screening
[edit]Date: 2014_9
Make miniprep, use Sal1 + EcoR1 to find positive clone
Cloning of 2321 pFastBacDual-VSVG-CAG-Flp
[edit]Purpose: Cloning of2321 pFastBacDual-VSVG-CAG-Flp-==
Date: 2014-9----
Notebook:0
Page:
Clone ID:2321
Experimenters: shanyi
Results
[edit]CLONING
[edit]PCR CAG-pH-T7 promoters from 298 pTriEx-HTNC
2364-CAG-Xho1-F atagcactcgagTATATGGAGTTCCGCGTTAC
2364-CAG-BamH1-R tgtcatggatccTATATCTCCTTTGATTGTAAATAAAATG
Digest with Xho1 + BamH1.
Digest vector with Xho1 + BamH1.
Colony Screening
[edit]Date: 2014_9
Make miniprep, use Xho1 + BamH1 to find positive clone
digest with NdeI+XhoI should get four bands,digest with XbaI should get two bands
send 2321-2 and 2321-3 to sequence with NLS-HindIII-R(NLS-HindIII-R accttcctcttcttcttcaagcttacatac), poor quality or no priming
use primers to do pcr screening
2364-CAG-Xho1-F atagcactcgagTATATGGAGTTCCGCGTTAC
hIgG-Lc-BamH1-Nhe1-B ctagcggccatggccggctgggccgcgagtaataacaatccagcggctgccgtaggcaataggtatttcatggtggcg
