LOCUS Exported 6087 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Scrambled control for Cre-dependent knockdown of Scn9a (Nav1.7). ACCESSION . VERSION . KEYWORDS AAV-FLEX-EGFP-mir30(Scn9a-scrambled) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6087) AUTHORS Branco T, Tozer A, Magnus CJ, Sugino K, Tanaka S, Lee AK, Wood JN, Sternson SM TITLE Near-Perfect Synaptic Integration by Nav1.7 in Hypothalamic Neurons Regulates Body Weight. JOURNAL Cell. 2016 Jun 16;165(7):1749-61. doi: 10.1016/j.cell.2016.05.019. PUBMED 27315482 REFERENCE 2 (bases 1 to 6087) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6087 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(11..28) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(182..770) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(262..281) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(941..1801) /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 1564..1583 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(1802..1906) /gene="bla" /label=AmpR promoter rep_origin complement(1932..2387) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2069..2090) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 2281..2300 /label=F1ori-R /note="F1 origin, reverse primer" repeat_region 2459..2588 /label=AAV2 ITR promoter 2901..3178 /label=chicken beta-actin promoter primer_bind 3458..3480 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 3542..3561 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" protein_bind 3597..3630 /label=loxN /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GTATACCT). loxN sites are compatible with each other, but incompatible with loxP or lox2272 sites." protein_bind 3597..3630 /label=lox2272 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GGATACTT). lox2272 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." protein_bind 3649..3682 /label=lox5171 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GTACACAT). lox5171 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." ncRNA complement(3753..3879) /label=3' miR-30a /note="sequence downstream of the 71-nt precursor of the human miR-30a microRNA (Zeng et al., 2002)" /ncRNA_class="miRNA" ncRNA complement(3907..3921) /label=miR-30a loop /note="loop from the the 71-nt precursor of the human miR-30a microRNA (Zeng et al., 2002)" /ncRNA_class="miRNA" ncRNA complement(3949..4070) /label=5' miR-30a /note="sequence upstream of the 71-nt precursor of the human miR-30a microRNA (Zeng et al., 2002)" /ncRNA_class="miRNA" CDS complement(4134..4853) /codon_start=1 /product="humanized Renilla green fluorescent protein" /label=hrGFP /note="mammalian codon-optimized" /translation="MVSKQILKNTGLQEIMSFKVNLEGVVNNHVFTMEGCGKGNILFGN QLVQIRVTKGAPLPFAFDILSPAFQYGNRTFTKYPEDISDFFIQSFPAGFVYERTLRYE DGGLVEIRSDINLIEEMFVYRVEYKGRNFPNDGPVMKKTITGLQPSFEVVYMNDGVLVG QVILVYRLNSGKFYSCHMRTLMKSKGVVKDFPEYHFIQHRLEKTYVEDGGFVEQHETAI AQLTSLGKPLGSLHEWV" regulatory 4850..4859 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" protein_bind complement(4864..4897) /label=loxN /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GTATACCT). loxN sites are compatible with each other, but incompatible with loxP or lox2272 sites." protein_bind complement(4864..4897) /label=lox2272 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GGATACTT). lox2272 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." protein_bind complement(4942..4975) /label=lox5171 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GTACACAT). lox5171 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." misc_feature 4989..5577 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(5042..5062) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(5460..5471) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 5620..5741 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5631..5650) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 5685..5704 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 5903..6032 /label=AAV2 ITR ORIGIN 1 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat 61 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga 121 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt 181 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt 241 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc 301 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa 361 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct 421 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta 481 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg 541 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc 601 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta 661 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg 721 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt 781 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg 841 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta 901 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg 961 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg 1021 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc 1081 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg 1141 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg 1201 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag 1261 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat 1321 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc 1381 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc 1441 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa 1501 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac 1561 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt 1621 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc 1681 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa 1741 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca 1801 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat 1861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa 1921 aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta 1981 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga 2041 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa 2101 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga 2161 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc 2221 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga 2281 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg 2341 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccatt 2401 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct 2461 gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg cgacctttgg 2521 tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact ccatcactag 2581 gggttccttg tagttaatga ttaacccgcc atgctactta tctacgtagc catgctctag 2641 gaagatcgta ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac 2701 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca 2761 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg 2821 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt 2881 agtcatcgct attaccatgg tcgaggtgag ccccacgttc tgcttcactc tccccatctc 2941 ccccccctcc ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat 3001 gggggcgggg gggggggggg ggcgcgcgcc aggcggggcg gggcggggcg aggggcgggg 3061 cggggcgagg cggagaggtg cggcggcagc caatcagagc ggcgcgctcc gaaagtttcc 3121 ttttatggcg aggcggcggc ggcggcggcc ctataaaaag cgaagcgcgc ggcgggcggg 3181 agtcgctgcg acgctgcctt cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc 3241 cccggctctg actgaccgcg ttactcccac aggtgagcgg gcgggacggc ccttctcctc 3301 cgggctgtaa ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg ctgcgtgaaa 3361 gccttgaggg gctccgggag ggccctttgt gcggggggag cggctcgggg ctgtccgcgg 3421 ggggacggct gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc 3481 ggcggctcta gagcctctgc taaccatgtt catgccttct tctttttcct acagctcctg 3541 ggcaacgtgc tggttattgt gctgtctcat cattttggca aagaattgga tcctccataa 3601 cttcgtatag gataccttat acgaagttat ctcagtttaa actaagccat aacttcgtat 3661 agtacacatt atacgaagtt attgtttacc ggtacctgga ggaaaaaaaa aaggggagag 3721 tcaggggtcg cgaattagtc ttccaattga aaaaagtgat ttaatttata ccattttaat 3781 tcagctttgt aaaaatgtat caaagagata gcaaggtatt cagttttagt aaacaagata 3841 attgctccta aagtagcccc ttgaattccg aggcagtagg cagcgaagtc agggaggaat 3901 ttattacatc tgtggcttca ctaataaatt cctccctgac ttcgtcgctc actgtcaaca 3961 gcaatatacc ttctcgagcc ttctgttggg ttaacctgaa gaagtaatcc cagcaagtgt 4021 ttccaagatg tgcaggcaac gattctgtaa agtactgaag cctcattcaa catagtatat 4081 gtgctgccga agcgagcact taacaaggct tgatatccac tcacctgcaa ttgttacacc 4141 cactcgtgca ggctgcccag gggcttgccc aggctggtca gctgggcgat ggcggtctcg 4201 tgctgctcca cgaagccgcc gtcctccacg taggtcttct ccaggcggtg ctggatgaag 4261 tggtactcgg ggaagtcctt caccacgccc ttgctcttca tcagggtgcg catgtggcag 4321 ctgtagaact tgccgctgtt caggcggtac accaggatca cctggcccac cagcacgccg 4381 tcgttcatgt acaccacctc gaagctgggc tgcaggccgg tgatggtctt cttcatcacg 4441 gggccgtcgt tggggaagtt gcggcccttg tactccacgc ggtacacgaa catctcctcg 4501 atcaggttga tgtcgctgcg gatctccacc aggccgccgt cctcgtagcg cagggtgcgc 4561 tcgtacacga agccggcggg gaagctctgg atgaagaagt cgctgatgtc ctcggggtac 4621 ttggtgaagg tgcggttgcc gtactggaag gcggggctca ggatgtcgaa ggcgaagggc 4681 aggggggcgc ccttggtcac gcggatctgc accagctggt tgccgaacag gatgttgccc 4741 ttgccgcagc cctccatggt gaacacgtgg ttgttcacca cgccctccag gttcaccttg 4801 aagctcatga tctcctgcag gccggtgttc ttcaggatct gcttgctcac catggtggct 4861 agcataactt cgtataaggt atcctatacg aagttatcca ttcaggctgt gtagcatcaa 4921 tggcatggca caaacttagc cataacttcg tataatgtgt actatacgaa gttatcaagc 4981 ttatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact 5041 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg 5101 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg 5161 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa 5221 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc 5281 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg 5341 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt 5401 ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt 5461 cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc 5521 cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc 5581 gataccgtcg acccgggcgg ccgcttcgag cagacatgat aagatacatt gatgagtttg 5641 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta 5701 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc 5761 attttatgtt tcaggttcag ggggagatgt gggaggtttt ttaaagcaag taaaacctct 5821 acaaatgtgg taaaatcgat aaggatcttc ctagagcatg gctacgtaga taagtagcat 5881 ggcgggttaa tcattaacta caaggaaccc ctagtgatgg agttggccac tccctctctg 5941 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc 6001 cgggcggcct cagtgagcga gcgagcgcgc agctgcatta atgaatcggc caacgcgcgg 6061 ggagaggcgg tttgcgtatt gggcgct //