LOCUS Exported 5755 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Reporter of botulinum toxin B cleavage. ACCESSION . VERSION . KEYWORDS pQL47 EGFP-VAMP-GST SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5755) AUTHORS Liu Q, Sinnen BL, Boxer EE, Schneider MW, Grybko MJ, Buchta WC, Gibson ES, Wysoczynski CL, Ford CP, Gottschalk A, Aoto J, Tucker CL, Kennedy MJ TITLE A Photoactivatable Botulinum Neurotoxin for Inducible Control of Neurotransmission. JOURNAL Neuron. 2019 Jan 15. pii: S0896-6273(19)30003-0. doi: 10.1016/j.neuron.2019.01.002. PUBMED 30704911 REFERENCE 2 (bases 1 to 5755) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5755 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 61..364 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 365..568 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 519..539 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 607..616 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 613..1329 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 613..1329 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(658..679) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(919..938) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 1266..1287 /label=EGFP-C /note="EGFP, forward primer" CDS 1759..2406 /codon_start=1 /product="glutathione S-transferase from Schistosoma japonicum" /label=GST /translation="PILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKKFE LGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRYGV SRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVLYM DPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK" primer_bind 2364..2386 /label=pGEX 5' /note="pGEX vectors, Glutathione-S-transferase, forward primer" polyA_signal 2543..2664 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2580..2599) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2634..2653 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(2671..3126) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2808..2829) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3020..3039 /label=F1ori-R /note="F1 origin, reverse primer" promoter 3153..3257 /gene="bla" /label=AmpR promoter promoter 3259..3616 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(3282..3302) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3467..3602 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3529..3548 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3651..4445 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(3705..3724) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4315..4334 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(4633..4652) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 4677..4724 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 5053..5641 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5542..5561 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg 61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt 121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca 181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc 241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta 301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac 361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg 421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg 481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt 541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta 601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg 661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc 721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg 781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc 841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag 901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag 961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac 1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac 1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc 1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg 1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc 1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag 1321 ctgtacaagt actcagatct cgagctcaag cttcgaattc tgcagtcgac ggtaccgagc 1381 tcggatccac tagtccagtg tggtggaatt gcccttatgt ctacaggtcc aactgctgcc 1441 actggcagta atcgaagact tcagcagaca caaaatcaag tagatgaggt ggtggacata 1501 atgcgagtta acgtggacaa ggttctggaa agagaccaga agctctctga gttagacgac 1561 cgtgcagacg cactgcaggc aggcgcttct caatttgaaa cgagcgcagc caagttgaag 1621 aggaaatatt ggtggaagaa ttgcaagatg tgggcaatcg ggattactgt tctggttatc 1681 ttcatcatca tcatcatcgt gtgggttgtc tcttccgcgg gtggaggttc aggaggaggc 1741 ggccgcatga ccaagttacc tatactaggt tattggaaaa ttaagggcct tgtgcaaccc 1801 actcgacttc ttttggaata tcttgaagaa aaatatgaag agcatttgta tgagcgcgat 1861 gaaggtgata aatggcgaaa caaaaagttt gaattgggtt tggagtttcc caatcttcct 1921 tattatattg atggtgatgt taaattaaca cagtctatgg ccatcatacg ttatatagct 1981 gacaagcaca acatgttggg tggttgtcca aaagagcgtg cagagatttc aatgcttgaa 2041 ggagcggttt tggatattag atacggtgtt tcgagaattg catatagtaa agactttgaa 2101 actctcaaag ttgattttct tagcaagcta cctgaaatgc tgaaaatgtt cgaagatcgt 2161 ttatgtcata aaacatattt aaatggtgat catgtaaccc atcctgactt catgttgtat 2221 gacgctcttg atgttgtttt atacatggac ccaatgtgcc tggatgcgtt cccaaaatta 2281 gtttgtttta aaaaacgtat tgaagctatc ccacaaattg ataagtactt gaaatccagc 2341 aagtatatag catggccttt gcagggctgg caagccacgt ttggtggtgg cgaccatcct 2401 ccaaaatcgg atcatctggt ttgagtctag ataactgatc ataatcagcc ataccacatt 2461 tgtagaggtt ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa 2521 aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag 2581 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt 2641 gtccaaactc atcaatgtat cttaacgcgt aaattgtaag cgttaatatt ttgttaaaat 2701 tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa 2761 tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca 2821 agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg 2881 gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta 2941 aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg 3001 cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa 3061 gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg 3121 gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct 3181 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat 3241 attgaaaaag gaagagtcct gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg 3301 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta 3361 gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat 3421 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac 3481 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga 3541 ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg 3601 cctaggcttt tgcaaagatc gatcaagaga caggatgagg atcgtttcgc atgattgaac 3661 aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact 3721 gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc 3781 gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg caagacgagg 3841 cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg 3901 tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt 3961 catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc 4021 atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag 4081 cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg 4141 ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc 4201 tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt 4261 ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg 4321 ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt 4381 acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct 4441 tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg 4501 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga 4561 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccctag 4621 ggggaggcta actgaaacac ggaaggagac aataccggaa ggaacccgcg ctatgacggc 4681 aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt gttcataaac gcggggttcg 4741 gtcccagggc tggcactctg tcgatacccc accgagaccc cattggggcc aatacgcccg 4801 cgtttcttcc ttttccccac cccacccccc aagttcgggt gaaggcccag ggctcgcagc 4861 caacgtcggg gcggcaggcc ctgccatagc ctcaggttac tcatatatac tttagattga 4921 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat 4981 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat 5041 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa 5101 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa 5161 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt 5221 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt 5281 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata 5341 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt 5401 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac 5461 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga 5521 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg 5581 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa 5641 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat 5701 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcca tgcat //