pOTB7D is derived from pOTB7 where a pair of oligos corresponding to
gaattccactgtgtggcggccgccaccctgtgctcgag were annealed and ligated into the
EcoRI and XhoI prepared pOTB7. This introduced two DraIII sites that allow
for directional cloning of cDNA inserts that contain SfiI sites. cDNA inserts
will have their 5' ends near the EcoRI site and the polyA tail/3' ends near
the XhoI site.
>pOTB7D polylinker:
SP6(52,72)>>>
|
1 CGTTAGAACGCGGCTACAATTAATACATAACCTTATGTATCATACACATACGATTTAGGT 60
GCAATCTTGCGCCGATGTTAATTATGTATTGGAATACATAGTATGTGTATGCTAAATCCA
BamHI M13 forw.(111,127)>>>
| |
61 GACACTATAGAACTCGAAGGATCCACAAGTTTGTACAAAAAAGCAGGCTTGTAAAACGAC 120
CTGTGATATCTTGAGCTTCCTAGGTGTTCAAACATGTTTTTTCGTCCGAACATTTTGCTG
AvaII StuI BglI EcoRI DraIII
| | | | |
121 GGCCAGTAACTATAACGGTCCTAAGGTAGCGAGGCCTGGGTGGCgaattccactgtgtgg 180
CCGGTCATTGATATTGCCAGGATTCCATCGCTCCGGACCCACCGCTTAAGGTGACACACC
DraIII XhoI
| |
181 cggccgccaccctgtgctcgagGCATCTATGTCGGGTGCGGAGAAAGAGGTAATGAAATG 240
GCCGGCGGTGGGACACGAGCTCCGTAGATACAGCCCACGCCTCTTTCTCCATTACTTTAC
M13 rev(244,262)<<< BglII
| |
241 GCACATGGTCATAGCTGTTTCCTGACCCAGCTTTCTTGTACAAAGTGGTAGATCTGCCGG 300
CGTGTACCAGTATCGACAAAGGACTGGGTCGAAAGAACATGTTTCACCATCTAGACGGCC
T7(304,323)<<< HpaI
| |
301 TCTCCCTATAGTGAGTCGTATTAATTTCGATAAGCCAGGTTAACCTGCATTAATGAATCG 360
AGAGGGATATCACTCAGCATAATTAAAGCTATTCGGTCCAATTGGACGTAATTACTTAGC
>pOTB7D 1827 bases.
CGTTAGAACGCGGCTACAATTAATACATAACCTTATGTATCATACACATACGATTTAGGT
GACACTATAGAACTCGAAGGATCCACAAGTTTGTACAAAAAAGCAGGCTTGTAAAACGAC
GGCCAGTAACTATAACGGTCCTAAGGTAGCGAGGCCTGGGTGGCgaattccactgtgtgg
cggccgccaccctgtgctcgagGCATCTATGTCGGGTGCGGAGAAAGAGGTAATGAAATG
GCACATGGTCATAGCTGTTTCCTGACCCAGCTTTCTTGTACAAAGTGGTAGATCTGCCGG
TCTCCCTATAGTGAGTCGTATTAATTTCGATAAGCCAGGTTAACCTGCATTAATGAATCG
GCTGCAGTACCCGGGAATTTAACCCGCCTAATGAGCGGGCTTTTTTTTGTGATCCAAAGG
ATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATCTGCTGCTTGCAAACAAAAAAACCACC
GCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAGCTACCAACTCTTTTTCCGAAGGTAAC
TGGCTTCAGCAGAGCGCAGATACCAAATACTGTTCTTCTAGTGTAGCCGTAGTTAGGCCA
CCACTTCAAGAACTCTGTAGCACCGCCTACATACCTCGCTCTGCTAATCCTGTTACCAGT
GGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACCGGGTTGGACTCAAGACGATAGTTACC
GGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGTTCGTGCACACAGCCCAGCTTGGAGCG
AACGACCTACACCGAACTGAGATACCTACAGCGTGAGCTATGAGAAAGCGCCACGCTTCC
CGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGCGGCAGGGTCGGAACAGGAGAGCGCAC
GAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCT
CTGACTTGAGCGTCGATTTTTGTGATGCTCGTCAGGGGGGCGGAGCCTATGGAAAAACGC
CAGCAACGCGGATCACAACAAAAAGCCCGCTCATTAGGCGGGCTAAATTCTCATGTTTGA
CAGCTTATCATCGATAAGCTAGCGGCCGCTAGCTTTAATGAGTTATCGAGATTTTCAGGA
GCTAAGGAAGCTAAAATGGAGAAAAAAATCACTGGATATACCACCGTTGATATATCCCAA
TGGCATCGTAAAGAACATTTTGAGGCATTTCAGTCAGTTGCTCAATGTACCTATAACCAG
ACCGTTCAGCTGGATATTACGGCCTTTTTAAAGACCGTAAAGAAAAATAAGCACAAGTTT
TATCCGGCCTTTATTCACATTCTTGCCCGCCTGATGAATGCTCATCCGGAGTTCCGTATG
GCAATGAAAGACGGTGAGCTGGTGATATGGGATAGTGTTCACCCTTGTTACACCGTTTTC
CATGAGCAAACTGAAACGTTTTCATCGCTCTGGAGTGAATACCACGACGATTTCCGGCAG
TTTCTACACATATATTCGCAAGATGTGGCGTGTTACGGTGAAAACCTGGCCTATTTCCCT
AAAGGGTTTATTGAGAATATGTTTTTCGTCTCAGCCAATCCCTGGGTGAGTTTCACCAGT
TTTGATTTAAACGTGGCCAATATGGACAACTTCTTCGCCCCCGTTTTCACCATGGGCAAA
TATTATACGCAAGGCGACAAGGTGCTGATGCCGCTGGCGATTCAGGTTCATCATGCCGTT
TGTGATGGCTTCCATGTCGGCAGAATGCTTAATGAATTACAACAGTACTGCGATGAGTGG
CAGGGCGGGGCGTAATTGGTACGTCGA
>pOTB7D with linkers; 1853 bases.
CGTTAGAACGCGGCTACAATTAATACATAACCTTATGTATCATACACATACGATTTAGGT
GACACTATAGAACTCGAAGGATCCACAAGTTTGTACAAAAAAGCAGGCTTGTAAAACGAC
GGCCAGTAACTATAACGGTCCTAAGGTAGCGAGGCCTGGGTGGCgaattccactgtcggc
cctgcgtttgctggctttgatgaaa[5'-cDNA-3']catgtcggccgcctgtgctcgagGCATCTATGTCG
GGTGCGGAGAAAGAGGTAATGAAATGGCACATGGTCATAGCTGTTTCCTGACCCAGCTTT
CTTGTACAAAGTGGTAGATCTGCCGGTCTCCCTATAGTGAGTCGTATTAATTTCGATAAG
CCAGGTTAACCTGCATTAATGAATCGGCTGCAGTACCCGGGAATTTAACCCGCCTAATGA
GCGGGCTTTTTTTTGTGATCCAAAGGATCTTCTTGAGATCCTTTTTTTCTGCGCGTAATC
TGCTGCTTGCAAACAAAAAAACCACCGCTACCAGCGGTGGTTTGTTTGCCGGATCAAGAG
CTACCAACTCTTTTTCCGAAGGTAACTGGCTTCAGCAGAGCGCAGATACCAAATACTGTT
CTTCTAGTGTAGCCGTAGTTAGGCCACCACTTCAAGAACTCTGTAGCACCGCCTACATAC
CTCGCTCTGCTAATCCTGTTACCAGTGGCTGCTGCCAGTGGCGATAAGTCGTGTCTTACC
GGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGGCTGAACGGGGGGT
TCGTGCACACAGCCCAGCTTGGAGCGAACGACCTACACCGAACTGAGATACCTACAGCGT
GAGCTATGAGAAAGCGCCACGCTTCCCGAAGGGAGAAAGGCGGACAGGTATCCGGTAAGC
GGCAGGGTCGGAACAGGAGAGCGCACGAGGGAGCTTCCAGGGGGAAACGCCTGGTATCTT
TATAGTCCTGTCGGGTTTCGCCACCTCTGACTTGAGCGTCGATTTTTGTGATGCTCGTCA
GGGGGGCGGAGCCTATGGAAAAACGCCAGCAACGCGGATCACAACAAAAAGCCCGCTCAT
TAGGCGGGCTAAATTCTCATGTTTGACAGCTTATCATCGATAAGCTAGCGGCCGCTAGCT
TTAATGAGTTATCGAGATTTTCAGGAGCTAAGGAAGCTAAAATGGAGAAAAAAATCACTG
GATATACCACCGTTGATATATCCCAATGGCATCGTAAAGAACATTTTGAGGCATTTCAGT
CAGTTGCTCAATGTACCTATAACCAGACCGTTCAGCTGGATATTACGGCCTTTTTAAAGA
CCGTAAAGAAAAATAAGCACAAGTTTTATCCGGCCTTTATTCACATTCTTGCCCGCCTGA
TGAATGCTCATCCGGAGTTCCGTATGGCAATGAAAGACGGTGAGCTGGTGATATGGGATA
GTGTTCACCCTTGTTACACCGTTTTCCATGAGCAAACTGAAACGTTTTCATCGCTCTGGA
GTGAATACCACGACGATTTCCGGCAGTTTCTACACATATATTCGCAAGATGTGGCGTGTT
ACGGTGAAAACCTGGCCTATTTCCCTAAAGGGTTTATTGAGAATATGTTTTTCGTCTCAG
CCAATCCCTGGGTGAGTTTCACCAGTTTTGATTTAAACGTGGCCAATATGGACAACTTCT
TCGCCCCCGTTTTCACCATGGGCAAATATTATACGCAAGGCGACAAGGTGCTGATGCCGC
TGGCGATTCAGGTTCATCATGCCGTTTGTGATGGCTTCCATGTCGGCAGAATGCTTAATG
AATTACAACAGTACTGCGATGAGTGGCAGGGCGGGGCGTAATTGGTACGTCGA