The Sekelsky Lab

pP{EndsOut2}

pP{EndsOut2} is a P element transformation vector for ends-out, or gene replacement, targeting. Genomic DNA is cloned into the polylinker between the I-SceI cut sites. A selectable marker, such as hsp70::white, is included within the fragment, either to disrupt a gene of interest, or cloned into a site that should not create a mutation (if, for example you want to make a site-directed mutation in the gene of interest). Gong and Golic reported successful ends-out targeting of yellow at the 2002 GSA Fly Meeting. We are currently attempting ends-out targeting of an autosomal locus.

pP{EndsOut2} replaces pP{EndsOut}, which was missing one of the I-SceI cut sites. In this version, the FRT sequences are immediately adjacent to the I-SceI cut sites, which are adjacent to the polylinker.

pP{EndsOut2} Sequence

CATGATGAAATAACATAAGGTGGTCCCGTCGGCAAGAGACATCCACTTAACGTATGCTT GCAATAAGTGCGAGTGAAAGGAATAGTATTCTGAGTGTCGTATTGAGTCTGAGTGAGAC AGCGATATGATTGTTGATTAACCCTTAGCATGTCCGTGGGGTTTGAATTAACTCATAAT ATTAATTAGACGAAATTATTTTTAAAGTTTTATTTTTAATAATTTGCGAGTACGCAAAG ctagagttgttggttggcacaccacaaatatactgttgccgagcacaattccgctcgac GAAGTTCCTATTCTCTAGAAAGTATAGGAACTTCTAGGGATAACAGGGTAATGGTACCC GCCCGGGGATCAGATCCGCGGCCGCACGCGTGAATTCGCTAGCATGCAGGCCTGCAGAT CTCGAGTAGGGATAACAGGGTAATGAAGTTCCTATTCTCTAGAAAGTATAGGAACTTCggatc caaacctagtaattaattcggctgctgctctAAACGACGCATTTCGTACTCCAAAGTAC GAATTTTTTCCCTCAAGCTCTTATTTTCATTAAACAATGAACAGGACCTAACGCACAGT CACGTTATTGTTTACATAAATGATTTTTTTTACTATTCAAACTTACTCTGTTTGTGTAC TCCCACTGGTATAGCCTTCTTTTATCTTTTCTGGTTCAGGCTCTATCACTTTACTAGGT ACGGCATCTGCGTTGAGTCGCCTCCTTTTAAATGTCTGACCTTTTGCAGGTGCAGCCTT CCACTGCGAATCATTAAAGTGGGTATCACAAATTTGGGAGTTTTCACCAAGGCTGCACC CAAGGCTCTGCTCCCACAATTTTCTCTTAATAGCACACTTCGGCACGTGAATTAATTTT ACTCCAGTCACAGCTTTGCAGCAAAATTTGCAATATTTCATTTTTTTTTATTCCACGTA AGGGTTAATGTTTTCAAAAAAAAATTCGTCCGCACACAACCTTTCCTCTCAACAAGCAA ACGTGCACTGAATTTAAGTGTATACTTCGGTAAGCTTCGGCTTTCGACGGGACCACCTT ATGTTATTTCATCATGggccagacccacgtagtccagcggcagatcggcggcggagaag ttaagcgtctccaggatgaccttgcccgaactggggcacgtggtgttcgacgatgtgca gctaatttcgcccggctccacgtccgcccattggttaatcagcagaccctcgttggcgt aacggaaccatgagaggtacgacaaccatttgaggtatactggcaccgagcccgagttc aagaagaaggcgtttttccataggctccgcccccctgacgagcatcacaaaaatcgacg ctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctg gaagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcc tttctcccttcgggaagcgtggcgctttctcatagctcacgctgtaggtatctcagttc ggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgacc gctgcgccttatccggtaactatcgtcttgagtccaacccggtaagacacgacttatcg ccactggcagcagccactggtaacaggattagcagagcgaggtatgtaggcggtgctac agagttcttgaagtggtggcctaactacggctacactagaaggacagtatttggtatct gcgctctgctgaagccagttaccttcggaaaaagagttggtagctcttgatccggcaaa caaaccaccgctggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaa aaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaacg aaaactcacgttaagggattttggtcatgagattatcaaaaaggatcttcacctagatc cttttaaattaaaaatgaagttttaaatcaatctaaagtatatatgagtaaacttggtc tgacagttaccaatgcttaatcagtgaggcacctatctcagcgatctgtctatttcgtt catccatagttgcctgactccccgtcgtgtagataactacgatacgggagggcttacca tctggccccagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatc agcaataaaccagccagccggaagggccgagcgcagaagtggtcctgcaactttatccg cctccatccagtctattaattgttgccgggaagctagagtaagtagttcgccagttaat agtttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgtttgg tatggcttcattcagctccggttcccaacgatcaaggcgagttacatgatcccccatgt tgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtcagaagtaagttggcc gcagtgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatc cgtaagatgcttttctgtgactggtgagtactcaaccaagtcattctgagaatagtgta tgcggcgaccgagttgctcttgcccggcgtcaatacgggataataccgcgccacatagc agaactttaaaagtgctcatcattggaaaacgttcttcggggcgaaaactctcaaggat cttaccgctgttgagatccagttcgatgtaacccactcgtgcacccaactgatcttcag catcttttactttcaccagcgtttctgggtgagcaaaaacaggaaggcaaaatgccgca aaaaagggaataagggcgacacggaaatgttgaatactcatactcttcctttttcaata ttattgaagcatttatcagggttattgtctcatgagcggatacatatttgaatgtattt agaaaaataaacaaataggggttccgcgcacatttccccgaaaagtgccacctgcgaag
tcttagagccagatatgcgagcacccggaagctcacgatgagaatggccagac

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