LOCUS Exported 7407 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Cre-conditional expression of the SthK channel. ACCESSION . VERSION . KEYWORDS pAAV-Syn-FLEX-Sthk-P2A-bPAC_minWPRE SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7407) AUTHORS Bernal Sierra YA, Rost BR, Pofahl M, Fernandes AM, Kopton RA, Moser S, Holtkamp D, Masala N, Beed P, Tukker JJ, Oldani S, Bonigk W, Kohl P, Baier H, Schneider-Warme F, Hegemann P, Beck H, Seifert R, Schmitz D TITLE Potassium channel-based optogenetic silencing. JOURNAL Nat Commun. 2018 Nov 5;9(1):4611. doi: 10.1038/s41467-018-07038-8. PUBMED 30397200 REFERENCE 2 (bases 1 to 7407) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7407 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..141 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 12..141 /label=AAV2 ITR promoter 173..620 /label=hSyn promoter /note="human synapsin I promoter; confers neuron-specific expression (Kugler et al., 2003)" protein_bind 635..668 /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 719..752 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." CDS complement(828..1535) /codon_start=1 /product="monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)" /label=mCherry /note="mammalian codon-optimized" /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE GRHSTGGMDELYK" primer_bind complement(1106..1125) /label=mCherry-F /note="mCherry, forward primer" primer_bind 1375..1393 /label=mCherry-R /note="mCherry, reverse primer" CDS complement(1551..1568) /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" CDS complement(1578..1607) /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" CDS complement(2697..2753) /codon_start=1 /product="2A peptide from porcine teschovirus-1 polyprotein" /label=P2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="ATNFSLLKQAGDVEENPGP" CDS complement(3455..3466) /codon_start=1 /product="WELQut protease recognition and cleavage site" /label=WELQut site /translation="WELQ" protein_bind complement(4095..4128) /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(4179..4212) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." primer_bind complement(4274..4294) /label=WPRE-R /note="WPRE, reverse primer" repeat_region 4670..4810 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 4670..4799 /label=AAV2 ITR rep_origin 4885..5340 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4972..4991) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 5182..5203 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(5357..5376) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 5476..5498 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(5536..5554) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 5622..5726 /gene="bla" /label=AmpR promoter CDS 5727..6587 /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(5945..5964) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 6758..7346 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 7247..7266 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc 61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca 121 actccatcac taggggttcc tgcggccgca cgcgtagagg gccctgcgta tgagtgcaag 181 tgggttttag gaccaggatg aggcggggtg ggggtgccta cctgacgacc gaccccgacc 241 cactggacaa gcacccaacc cccattcccc aaattgcgca tcccctatca gagaggggga 301 ggggaaacag gatgcggcga ggcgcgtgcg cactgccagc ttcagcaccg cggacagtgc 361 cttcgccccc gcctggcggc gcgcgccacc gccgcctcag cactgaaggc gcgctgacgt 421 cactcgccgg tcccccgcaa actccccttc ccggccacct tggtcgcgtc cgcgccgccg 481 ccggcccagc cggaccgcac cacgcgaggc gcgagatagg ggggcacggg cgcgaccatc 541 tgcgctgcgg cgccggcgac tcagcgctgc ctcagtctgc ggtgggcagc ggaggagtcg 601 tgtcgtgcct gagagcgcag tcgctagtac cggtataact tcgtatagga tactttatac 661 gaagttatgc agaatggtag ctggattgta gctgctatta gcaatatgaa acctcttaat 721 aacttcgtat agcatacatt atacgaagtt atttcgaact cgagccagag ctctggattt 781 aaatggcgcg cctattaatt aaggccgcga ctctagaatt cgctagctta cttgtacagc 841 tcgtccatgc cgccggtgga gtggcggccc tcggcgcgtt cgtactgttc cacgatggtg 901 tagtcctcgt tgtgggaggt gatgtccaac ttgatgttga cgttgtaggc gccgggcagc 961 tgcacgggct tcttggcctt gtaggtggtc ttgacctcag cgtcgtagtg gccgccgtcc 1021 ttcagcttca gcctctgctt gatctcgccc ttcagggcgc cgtcctcggg gtacatccgc 1081 tcggaggagg cctcccagcc catggtcttc ttctgcatta cggggccgtc ggaggggaag 1141 ttggtgccgc gcagcttcac cttgtagatg aactcgccgt cctgcaggga ggagtcctgg 1201 gtcacggtca ccacgccgcc gtcctcgaag ttcatcacgc gctcccactt gaagccctcg 1261 gggaaggaca gcttcaagta gtcggggatg tcggcggggt gcttcacgta ggccttggag 1321 ccgtacatga actgagggga caggatgtcc caggcgaagg gcagggggcc acccttggtc 1381 accttcagct tggcggtctg ggtgccctcg taggggcggc cctcgccctc gccctcgatc 1441 tcgaactcgt ggccgttcac ggagccctcc atgtgcacct tgaagcgcat gaactccttg 1501 atgatggcca tgttatcctc ctcgcccttg ctcacggcag atctagctgc atggtggtga 1561 tggtggtgcg atcccttcag gtcctcctcc gagatcagct tttgctcgta cgtccgcggc 1621 ttgtcgtttt ccagggtctg ccggatcagc tgggtgatct gcagggtgtc gtagtacttc 1681 tgggcctcat tcacggtgta gacctcgatg gcttcctgct tgcccttgac ctggtgggct 1741 cccaggttga tgaaggtcca ctgggcctgg cagcacttct tcacgccggc tgtaaaggcc 1801 agggcgtagg gcagctgtct tgtcagggct tccagcctgg cggccacgtt cacggcgtcc 1861 cccagcaggg tgtggtccat cttcaggctg ctgcccatgt tgccctcgat cacgtggccg 1921 tagctcaggc cgatgccggt gtacagcagg tgcagggggt tggtggcctc cacgtggtgc 1981 cgcagctgct tcagctcgga gatgatgtcc aggctggtcc ggatggcggc gtcgccctgc 2041 tctttggtga agctggccat cacgcagtcg ccgatgaact tggtcacttc gccgccgtag 2101 gcgctgatga tccgggtgca gatgctgaag taccggttga ccaggatgac cacctcgttc 2161 acgggcagct tctcggtcag ggtgctgaag gccaggatgt cgctgaagaa gatgatcttc 2221 tcgaccagct ggggctccac ggtcaggggg ttgatgccct ggttgatcag gtagatcact 2281 ctggcgggca tgtacttttc cagcacccgg tggctctggg tgatggtctg cagcaggctc 2341 ttgatgggct ggatcatcag ctcgctgttc tcgttcaggt tgatggtttt catggcccag 2401 ttggggaaca tccggtcggt gatgtcgtac tcggttttca ggcacaggat gttggtgtgc 2461 cggtcgtcca ccaggatttt cttgtacagc ttgtccacct tctcgttctc gccttccagg 2521 atctggaaga acaggccctg caggtacagc agcacgccgg tgatgttgtc ccgctggttg 2581 ttcttgatgg acaccttgcc gatccgctgg atctcttcca ggctcaggtg gccgctgatc 2641 ttgctgatgt acaccagccg cttcatcatg ctcgagaggc cttgaattcc agatctaggt 2701 ccagggttct cctccacgtc tccagcctgc ttcagcaggc tgaagttagt agctccgctt 2761 ccgcatgcga gctccttaag cccgggaccg gttccccgcc gtgatgtccc cccttccagt 2821 tcttctttcc tccgcacagc cagttcctga atctgggctg caatctcagg atagcgagac 2881 aggattctgt caaaggtttc tttatccagt ctgtacaggt cacagaaagt ccgtgctctc 2941 acagtagctg ttcgtggtgc tcgcagaatc agtgccattt ctccaaagaa ctggccctca 3001 gacaggatgg cataccgggt cttttcgtct gcggacagga cctccacgct gcctctgttg 3061 ataaagtaga catcgctgcc cagctcgcca gccctaatga tatattcccc aggtccgtag 3121 atcacgggct ccagatgcag aatgatatcg cggataaagt cttcgccggc ccctttgaac 3181 agtgggacct tctcaatcac gtcgccatgg atttccattg cgacagccag tcgcagtggg 3241 tgaggcagtt ccttcagcac ctcgcgttcc tcatacccgc gccgagtctc ccacagataa 3301 tcaaagtatt ccagaattct cctctgcagc tcaggtgaga ttttcttgta actcaggaaa 3361 gctgtcaccc gttccatcct ctctcggtgc agcagtttag cggcgtccag cttactgacc 3421 agagatgcaa tattcccgat gaccagtcca tacattgcag ctcccagcag ctcaatgacg 3481 atggtgtaca cggtctgaat gggtgtggat ggggtaatat ctccgtagcc gatggtagtc 3541 agtgtggtga ttgtccagta gaaggcgctc aggtatctgg ttcctgctgg ggactcgcta 3601 ggtggctgca gtgacatcca gccgcaggca atcccatggg ctgccagcag gatgaatccc 3661 accagactca gcagccgcag cagtgctggg ttaattctgt aagacaggat tcttgtagca 3721 gacctctgga cggagatcag cttcagcagc ctcaccaggg acagcaggct caggggtgat 3781 ggcaggtgca gggcgaacac cagcaggtcc agtggcagag cggccagcag atctggcagt 3841 cggctaggag ccagcagagt tggtttcctg ccgcttgtct ttcgtgactc aaaggcgaaa 3901 tccagaggaa tgtctgcgat cagagccaga ctggccagga tatccagagg cagcagggga 3961 ctaaacagag aggggaacac cagggtcagt gggattcgaa tagcgtaata cagagtgaca 4021 gccagaatgc cgactttcca gaccagggtg taggtggggt gggagaaggc ggagcttttc 4081 atggtggcgg atccataact tcgtataaag tatcctatac gaagttattt gccttaaccc 4141 agaaattatc actgttattc tttagaatgg tgcaaagaat aacttcgtat aatgtatgct 4201 atacgaagtt atgtcgagat aatcaacctc tggattacaa aatttgtgaa agattgactg 4261 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt 4321 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttag 4381 ttcttgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc 4441 tgttgggcac tgacaattcc gtggtgttta tttgtgaaat ttgtgatgct attgctttat 4501 ttgtaaccat tctagcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat 4561 tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca 4621 gggggagatg tgggaggttt tttaaagcac gtgcggaccg agcggccgca ggaaccccta 4681 gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca 4741 aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc 4801 tgcctgcagg ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac 4861 cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg 4921 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg 4981 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg 5041 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt 5101 tgggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt 5161 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta 5221 tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa 5281 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt 5341 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc 5401 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac 5461 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac 5521 gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa 5581 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt 5641 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc 5701 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc 5761 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa 5821 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg 5881 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag 5941 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc 6001 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta 6061 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg 6121 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca 6181 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac 6241 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat 6301 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg 6361 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata 6421 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta 6481 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa 6541 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag 6601 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg 6661 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact 6721 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg 6781 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc 6841 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata 6901 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta 6961 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc 7021 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg 7081 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac 7141 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg 7201 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt 7261 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct 7321 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg 7381 ccttttgctg gccttttgct cacatgt //