LOCUS Exported 6646 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Cre-dependent expression of Flp recombinase.. ACCESSION . VERSION . KEYWORDS pAAV-nef-CIAO2-Flp SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6646) AUTHORS Fischer KB, Collins HK, Callaway EM TITLE Sources of off-target expression from recombinase-dependent AAV vectors and mitigation with cross-over insensitive ATG-out vectors. JOURNAL Proc Natl Acad Sci U S A. 2019 Dec 16. pii: 1915974116. doi: 10.1073/pnas.1915974116. PUBMED 31843925 REFERENCE 2 (bases 1 to 6646) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6646 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..130 /label=AAV2 ITR (alternate) /note="Functional equivalent of wild-type AAV2 ITR" promoter 177..388 /label=EF-1-alpha core promoter /note="core promoter for human elongation factor EF-1-alpha" LTR 401..669 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from human T-cell leukemia virus (HTLV) type 1" regulatory 700..709 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" intron 1265..1330 /label=small t intron /note="SV40 (simian virus 40) small t antigen intron" polyA_signal 1908..2042 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1945..1964) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 1999..2018 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" polyA_signal 3393..3869 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(3592..3611) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 3909..4049 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 3909..4038 /label=AAV2 ITR rep_origin 4124..4579 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4211..4230) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4421..4442 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4596..4615) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 4715..4737 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(4775..4793) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 4861..4965 /gene="bla" /label=AmpR promoter CDS 4966..5826 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind complement(5184..5203) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5997..6585 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6486..6505 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt 61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact 121 aggggttcct gcggccgcac gcgtaaggat ctgcgatcgc tccggtgccc gtcagtgggc 181 agagcgcaca tcgcccacag tccccgagaa gttgggggga ggggtcggca attgaacggg 241 tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat gtcgtgtact ggctccgcct 301 ttttcccgag ggtgggggag aaccgtatat aagtgcagta gtcgccgtga acgttctttt 361 tcgcaacggg tttgccgcca gaacacagct gaagcttcga ggggctcgca tctctccttc 421 acgcgcccgc cgccctacct gaggccgcca tccacgccgg ttgagtcgcg ttctgccgcc 481 tcccgcctgt ggtgcctcct gaactgcgtc cgccgtctag gtaagtttaa agctcaggtc 541 gagaccgggc ctttgtccgg cgctcccttg gagcctacct agactcagcc ggctctccac 601 gctttgcctg accctgcttg ctcaactcta cgtctttgtt tcgttttctg ttctgcgccg 661 ttacagatcc aagctgtgac cggcgcctac ggatccgccg ccaccatggg aatgcatgct 721 atagcataca ttatacgaag ttatgagctc tcggggacac caaatatggc gatctcggcc 781 ttttcgtttc ttggagctgg gacatgtttg ccatcgatcc atctaccacc agaacggccg 841 ttagatctgc tgccaccgtt gtttccaccg aagaaaccac cgttgccgta accaccacga 901 cggttgttgc taaagaagct gccaccgcca cggccaccgt tgtagccgcc gttgttgtta 961 ttgtagttgc tactgttatt tctggcactt cttggttttc ctcttaagtg aggaggaaca 1021 taaccattct cgttgttgtc gttgatgctt aaattttgca cttgttcgct cagttcagcc 1081 ataatatgaa atgcttttct tttgttctta cggaatacca cttgccacct atcaccacaa 1141 ctaacttttt cccgttcctc catctctttt atattttttt tctcgaggga tctttgtgaa 1201 ggaaccttac ttctgtggtg tgacataatt ggacaaacta cctacagaga tttaaagctc 1261 taaggtaaat ataaaatttt taagtgtata atgtgttaaa ctactgattc taattgtttg 1321 tgtattttag attccaacct atggaactga tgaatgggag cagtggtgga atgcctttaa 1381 tgaggaaaac ctgttttgct cagaagaaat gccatctagt gatgatgagg ctactgctga 1441 ctctcaacat tctactcctc caaaaaagaa gaagagaaag gtagaagacc ccaaggactt 1501 tccttcagaa ttgctaagtt ttttgagtca tgctgtgttt agtaatagaa ctcttgcttg 1561 ctttgctatt tacaccacaa aggaaaaagc tgcactgcta tacaagaaaa ttatggaaaa 1621 atattctgta acctttataa gtaggcataa cagttataat cataacatac tgttttttct 1681 tactccacac aggcatagag tgtctgctat taataactat gctcaaaaat tgtgtacctt 1741 tagcttttta atttgtaaag gggttaataa ggaatatttg atgtatagtg ccttgactag 1801 agatcataat cagccatacc acatttgtag aggttttact tgctttaaaa aacctcccac 1861 acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac ttgtttattg 1921 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt 1981 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga 2041 tctgacatgg taagtgaatt cataacttcg tatagcatac attataggta ccgaggctag 2101 cagccagttc gacatcctgt gcaagacccc ccccaaggtg ctggtgcggc agttcgtgga 2161 gagattcgag aggcccagcg gcgagaagat cgccagctgt gccgccgagc tgacctacct 2221 gtgctggatg atcacccaca acggcaccgc catcaagagg gccaccttca tgagctacaa 2281 caccatcatc agcaacagcc tgagcttcga catcgtgaac aagagcctgc agttcaagta 2341 caagacccag aaggccacca tcctggaggc cagcctgaag aagctgatcc ccgcctggga 2401 gttcaccatc atcccttaca acggccagaa gcaccagagc gacatcaccg acatcgtgtc 2461 cagcctgcag ctgcagttcg agagcagcga ggaggccgac aagggcaaca gccacagcaa 2521 gaagatgctg aaggccctgc tgtccgaggg cgagagcatc tgggagatca ccgagaagat 2581 cctgaacagc ttcgagtaca ccagcaggtt caccaagacc aagaccctgt accagttcct 2641 gttcctggcc acattcatca actgcggcag gttcagcgac atcaagaacg tggaccccaa 2701 gagcttcaag ctggtgcaga acaagtacct gggcgtgatc attcagtgcc tggtgaccga 2761 gaccaagaca agcgtgtcca ggcacatcta ctttttcagc gccagaggca ggatcgaccc 2821 cctggtgtac ctggacgagt tcctgaggaa cagcgagccc gtgctgaaga gagtgaacag 2881 gaccggcaac agcagcagca acaagcagga gtaccagctg ctgaaggaca acctggtgcg 2941 cagctacaac aaggccctga agaagaacgc cccctacccc atcttcgcta tcaagaacgg 3001 ccctaagagc cacatcggca ggcacctgat gaccagcttt ctgagcatga agggcctgac 3061 cgagctgaca aacgtggtgg gcaactggag cgacaagagg gcctccgccg tggccaggac 3121 cacctacacc caccagatca ccgccatccc cgaccactac ttcgccctgg tgtccaggta 3181 ctacgcctac gaccccatca gcaaggagat gatcgccctg aaggacgaga ccaaccccat 3241 cgaggagtgg cagcacatcg agcagctgaa gggcagcgcc gagggcagca tcagataccc 3301 cgcctggaac ggcatcatca gccaggaggt gctggactac ctgagcagct acatcaacag 3361 gcggatctga ggcgcgccct cgagagatct acgggtggca tccctgtgac ccctccccag 3421 tgcctctcct ggccctggaa gttgccactc cagtgcccac cagccttgtc ctaataaaat 3481 taagttgcat cattttgtct gactaggtgt ccttctataa tattatgggg tggagggggg 3541 tggtatggag caaggggcaa gttgggaaga caacctgtag ggcctgcggg gtctattggg 3601 aaccaagctg gagtgcagtg gcacaatctt ggctcactgc aatctccgcc tcctgggttc 3661 aagcgattct cctgcctcag cctcccgagt tgttgggatt ccaggcatgc atgaccaggc 3721 tcagctaatt tttgtttttt tggtagagac ggggtttcac catattggcc aggctggtct 3781 ccaactccta atctcaggtg atctacccac cttggcctcc caaattgctg ggattacagg 3841 cgtgaaccac tgctcccttc cctgtccttc tgattttgta ggtaaccacg tgcggaccga 3901 gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct 3961 cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt 4021 gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca 4081 tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc 4141 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc 4201 ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc 4261 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc 4321 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg 4381 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact 4441 ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt 4501 tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa 4561 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag 4621 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc 4681 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt 4741 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag 4801 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg 4861 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga 4921 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat 4981 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca 5041 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc 5101 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca 5161 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg 5221 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca 5281 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata 5341 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag 5401 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg 5461 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca 5521 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta 5581 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct 5641 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca 5701 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag 5761 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat 5821 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt 5881 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa 5941 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga 6001 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg 6061 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc 6121 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag 6181 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc 6241 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg 6301 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac 6361 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga 6421 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt 6481 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag 6541 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg 6601 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgt //