LOCUS Exported 7029 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION produces AAV expressing Cal-light TM-CaM-NES-TEV-N-AsLOV2-TEVseq-tTA. ACCESSION . VERSION . KEYWORDS pAAV-TM-CaM-NES-TEV-N-AsLOV2-TEVseq-tTA SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7029) AUTHORS Lee D, Hyun JH, Jung K, Hannan P, Kwon HB TITLE A calcium- and light-gated switch to induce gene expression in activated neurons. JOURNAL Nat Biotechnol. 2017 Jun 26. doi: 10.1038/nbt.3902. PUBMED 28650460 REFERENCE 2 (bases 1 to 7029) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7029 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 6..453 /label=hSyn promoter /note="human synapsin I promoter; confers neuron-specific expression (Kugler et al., 2003)" promoter 474..492 /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind 474..491 /label=SP6 /note="SP6 promoter, forward primer" sig_peptide 511..573 /product="leader sequence from mouse immunoglobulin kappa light chain" /label=Ig-kappa leader CDS 574..603 /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" CDS 607..753 /codon_start=1 /product="transmembrane domain from platelet derived growth factor receptor beta" /label=PDGFR-beta TM domain /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ KKPR" CDS 1294..1320 /codon_start=1 /product="nuclear export signal from the HIV Rev protein (Fischer et al., 1995)" /label=NES /translation="LPPLERLTL" CDS 2113..2133 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQG" CDS 2134..3138 /codon_start=1 /product="tetracycline-controlled transactivator, comprising a fusion of the tetracycline repressor TetR with the C-terminal activation domain of herpes simplex virus VP16" /label=tTA /note="In the Tet-Off(R) system, tTA binds to the Tet-responsive element and stimulates transcription only in the absence of tetracycline or doxycycline." /translation="SRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHVK NKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTRP TEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTTD SMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTIE GLLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHAD ALDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG" primer_bind complement(2204..2223) /label=Tet-R /note="Tetracycline resistance gene, reverse primer" misc_feature 3157..3745 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(3210..3230) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3628..3639) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3786..3993 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" repeat_region 4163..4292 /label=AAV2 ITR primer_bind complement(4358..4375) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4529..5117) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4609..4628) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5288..6148) /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 5911..5930 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6149..6253) /gene="bla" /label=AmpR promoter rep_origin complement(6279..6734) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(6416..6437) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 6628..6647 /label=F1ori-R /note="F1 origin, reverse primer" repeat_region 6806..6935 /label=AAV2 ITR ORIGIN 1 gtatgagtgc aagtgggttt taggaccagg atgaggcggg gtgggggtgc ctacctgacg 61 accgaccccg acccactgga caagcaccca acccccattc cccaaattgc gcatccccta 121 tcagagaggg ggaggggaaa caggatgcgg cgaggcgcgt gcgcactgcc agcttcagca 181 ccgcggacag tgccttcgcc cccgcctggc ggcgcgcgcc accgccgcct cagcactgaa 241 ggcgcgctga cgtcactcgc cggtcccccg caaactcccc ttcccggcca ccttggtcgc 301 gtccgcgccg ccgccggccc agccggaccg caccacgcga ggcgcgagat aggggggcac 361 gggcgcgacc atctgcgctg cggcgccggc gactcagcgc tgcctcagtc tgcggtgggc 421 agcggaggag tcgtgtcgtg cctgagagcg cagtcgagag gatctaagct ttcatttagg 481 tgacactata gaatacaagc tacttgttct atggagacag acacactcct gctatgggta 541 ctgctgctct gggttccagg ttccactggt gacgaacaaa aactcatctc agaagaggat 601 ctgaatgctg tgggccagga cacgcaggag gtcatcgtgg tgccacactc cttgcccttt 661 aaggtggtgg tgatctcagc catcctggcc ctggtggtgc tcaccatcat ctcccttatc 721 atcctcatca tgctttggca gaagaagcca cgtggtggct ctggaggtct cgagatgccg 781 gaccaactga ctgaagagca gatcgcagaa tttaaagagg ctttctccct atttgacaag 841 gacggggatg ggacaataac aaccaaggag ctggggacgg tgatgcggtc tctggggcag 901 aaccccacag aagcagagct gcaagacatg atcaatgaag tagatgccga cggtgacggc 961 acaatcgact tccctgagtt cctgacaatg atggcaagaa aaatgaaata cagggacacg 1021 gaagaagaaa ttagagaagc gttcggtgtg tttgataagg atggcaatgg ctacatcagt 1081 gcagcagagc ttcgccacgt gatgacaaac cttggagaga agttaacaga tgaagaggtt 1141 gatgaaatga tcagggaagc agacatcgat ggggatggtc aggtaaacta cgaagagttt 1201 gtacaaatga tgacagcgaa ggcatgcggt ggcggtggct ctggaggtgg tgggtccgga 1261 ggaggcggcc gctccggtgg atccatgctt caacttcctc ctcttgaacg tcttactctc 1321 gagatgggcg agagcctgtt caagggacct agggactaca accctatcag tagcacaatt 1381 tgtcacctga ccaacgagag tgatggccac acaacaagcc tgtacggcat cgggttcgga 1441 ccctttatca tcaccaacaa gcacctgttc aggcggaata atggcactct gctggtgcag 1501 agcctgcacg gggtgttcaa agtgaagaac acaaccactc tgcagcagca cctgatcgat 1561 gggcgggata tgatcatcat taggatgccc aaggacttcc ccccttttcc tcagaaactg 1621 aagttccgag agccccagag agaggagaga atctgtctgg tgaccacaaa ctttcaggag 1681 ctcggtagtg gtagtgggga gtttctggca accacactgg aacggatcga gaaaaatttc 1741 gtgattactg atccgagact gcctgacaac ccaatcattt ttgcgagcga ttccttcctg 1801 cagctgacag aatattctcg ggaagagatc ctggggcgca attgccgttt tctgcaggga 1861 cccgagacag accgtgccac tgttcggaaa atcagagatg ctattgacaa ccagactgaa 1921 gtgaccgttc agctgatcaa ttataccaag agcggcaaga agttctggaa cgtgttccac 1981 ctgcagccga tgcgcgatta taagggcgac gtccagtact tcattggcgt gcagctggat 2041 ggcaccgaac gtcttcatgg cgccgctgag cgtgaggcgg tctgcctgat caaaaagaca 2101 gcctttcaga ttgagaacct gtacttccag ggctctagat tagataaaag taaagtgatt 2161 aacagcgcat tagagctgct taatgaggtc ggaatcgaag gtttaacaac ccgtaaactc 2221 gcccagaagc taggtgtaga gcagcctaca ttgtattggc atgtaaaaaa taagcgggct 2281 ttgctcgacg ccttagccat tgagatgtta gataggcacc atactcactt ttgcccttta 2341 gaaggggaaa gctggcaaga ttttttacgt aataacgcta aaagttttag atgtgcttta 2401 ctaagtcatc gcgatggagc aaaagtacat ttaggtacac ggcctacaga aaaacagtat 2461 gaaactctcg aaaatcaatt agccttttta tgccaacaag gtttttcact agagaatgca 2521 ttatatgcac tcagcgctgt ggggcatttt actttaggtt gcgtattgga agatcaagag 2581 catcaagtcg ctaaagaaga aagggaaaca cctactactg atagtatgcc gccattatta 2641 cgacaagcta tcgaattatt tgatcaccaa ggtgcagagc cagccttctt attcggcctt 2701 gaattgatca tatgcggatt agaaaaacaa cttaaatgtg aaagtgggtc cgcgtacagc 2761 cgcgcgcgta cgaaaaacaa ttacgggtct accatcgagg gcctgctcga tctcccggac 2821 gacgacgccc ccgaagaggc ggggctggcg gctccgcgcc tgtcctttct ccccgcggga 2881 cacacgcgca gactgtcgac ggcccccccg accgatgtca gcctggggga cgagctccac 2941 ttagacggcg aggacgtggc gatggcgcat gccgacgcgc tagacgattt cgatctggac 3001 atgttggggg acggggattc cccgggtccg ggatttaccc cccacgactc cgccccctac 3061 ggcgctctgg atatggccga cttcgagttt gagcagatgt ttaccgatgc ccttggaatt 3121 gacgagtacg gtgggtaggc tagccagctt atcgataatc aacctctgga ttacaaaatt 3181 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct 3241 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg 3301 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc 3361 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt 3421 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc 3481 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg 3541 ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct gtgttgccac ctggattctg 3601 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc 3661 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg 3721 atctcccttt gggccgcctc cccgcatcga taccgtcgac ccgggcggcc gctcataaag 3781 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt 3841 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca 3901 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga 3961 ggattgggaa gacaatagca ggcatgctgg ggatgcggtc cggactgtac aattgcattc 4021 attttatgtt tcaggttcag ggggagatgt gggaggtttt ttaaagcaag taaaacctct 4081 acaaatgtgg taaaatcgat aaggatcttc ctagagcatg gctacgtaga taagtagcat 4141 ggcgggttaa tcattaacta caaggaaccc ctagtgatgg agttggccac tccctctctg 4201 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc 4261 cgggcggcct cagtgagcga gcgagcgcgc agctgcatta atgaatcggc caacgcgcgg 4321 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct 4381 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca 4441 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga 4501 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc 4561 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg 4621 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat 4681 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt 4741 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc 4801 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg 4861 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg 4921 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg 4981 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg 5041 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca 5101 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga 5161 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga 5221 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt 5281 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt 5341 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat 5401 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag 5461 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct 5521 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt 5581 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg 5641 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca 5701 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt 5761 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat 5821 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac 5881 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa 5941 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt 6001 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt 6061 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa 6121 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt 6181 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa 6241 taggggttcc gcgcacattt ccccgaaaag tgccacctaa attgtaagcg ttaatatttt 6301 gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat 6361 cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt 6421 ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt 6481 ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag 6541 gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg 6601 aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc 6661 gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc 6721 gctacagggc gcgtcccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt 6781 gcgggcctct tcgctattac gccagctgcg cgctcgctcg ctcactgagg ccgcccgggc 6841 aaagcccggg cgtcgggcga cctttggtcg cccggcctca gtgagcgagc gagcgcgcag 6901 agagggagtg gccaactcca tcactagggg ttccttgtag ttaatgatta acccgccatg 6961 ctacttatct acgtagccat gctctaggaa gatcgtaggt taattaatct agactgcaga 7021 gggccctgc //