LOCUS Exported 4777 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expresses the proton pump rhodopsin in mammalian cells under CMV promoter. ACCESSION . VERSION . KEYWORDS pN3-pHluorinCAAX SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4777) AUTHORS Wu L, Dong A, Dong L, Wang SQ, Li Y TITLE PARIS, an optogenetic method for functionally mapping gap junctions. JOURNAL Elife. 2019 Jan 14;8. pii: 43366. doi: 10.7554/eLife.43366. PUBMED 30638447 REFERENCE 2 (bases 1 to 4777) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4777 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 109..412 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 413..616 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 567..587 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" misc_feature 657..713 /label=MCS /note="multiple cloning site" regulatory 717..726 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 723..1439 /codon_start=1 /product="pH-sensitive mutant of green fluorescent protein (Ng et al., 2002)" /label=superecliptic pHluorin /note="excited strongly by blue light at neutral pH but minimally fluorescent at acidic pH" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNDHQVYIMADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLFTTSTLSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" primer_bind complement(756..784) /label=GFP-R /note="GFP, reverse primer. Does NOT anneal to EGFP" primer_bind 1379..1399 /label=GFP-F /note="GFP, forward primer. Does NOT anneal to EGFP" polyA_signal 1613..1734 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1650..1669) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 1704..1723 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(1741..2196) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(1878..1899) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 2090..2109 /label=F1ori-R /note="F1 origin, reverse primer" promoter 2223..2327 /gene="bla" /label=AmpR promoter promoter 2329..2686 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(2352..2372) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 2537..2672 /label=SV40 ori /note="SV40 origin of replication" primer_bind 2599..2618 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 2721..3515 /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(2775..2794) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 3385..3404 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(3703..3722) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 3747..3794 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 4123..4711 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4612..4631 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 cctgcgttat cccctgattc tgtggataac cgtattaccg ccatgcatta gttattaata 61 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact 121 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat 181 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta 241 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc 301 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg 361 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg 421 gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct 481 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa 541 atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt 601 ctatataagc agagctggtt tagtgaaccg tcagatccgc tagcgctacc ggactcagat 661 ctcgagctca agcttcgaat tctgcagtcg acggtaccgc gggcccggga tccatcgcca 721 ccatggtgag taaaggagaa gaacttttca ctggagttgt cccaattctt gttgaattag 781 atggtgatgt taatgggcac aaattttctg tcagtggaga gggtgaaggt gatgcaacat 841 acggaaaact tacccttaaa tttatttgca ctactggaaa actacctgtt ccttggccaa 901 cacttgtcac tactttaact tatggtgttc aatgcttttc aagataccca gatcatatga 961 aacggcatga ctttttcaag agtgccatgc ccgaaggtta tgtacaggaa agaactatat 1021 ttttcaaaga tgacgggaac tacaagacac gtgctgaagt caagtttgaa ggtgataccc 1081 ttgttaatag aatcgagtta aaaggtattg attttaaaga agatggaaac attcttggac 1141 acaaattgga atacaactat aacgatcacc aggtgtacat catggcagac aaacaaaaga 1201 atggaatcaa agctaacttc aaaattagac acaacattga agatggaggc gttcaactag 1261 cagaccatta tcaacaaaat actccaattg gcgatgggcc cgtcctttta ccagacaacc 1321 attacctgtt tacaacttct actctttcga aagatcccaa cgaaaagaga gaccacatgg 1381 tccttcttga gtttgtaaca gctgctggga ttacacatgg catggatgaa ctatacaaag 1441 atggtaaaaa gaagaaaaag aagtcaaaga caaagtgtgt aattatgtaa agcggccgcg 1501 actctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct 1561 cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt 1621 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc 1681 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaaggcgt 1741 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat 1801 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga 1861 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca 1921 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct 1981 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc 2041 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag 2101 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca 2161 cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg 2221 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga 2281 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa 2341 gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg 2401 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg 2461 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc 2521 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca 2581 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt 2641 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga 2701 caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg 2761 cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg 2821 ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt 2881 ccggtgccct gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg 2941 gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat 3001 tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat 3061 ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg 3121 accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg 3181 atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc 3241 tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc 3301 cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg 3361 tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg 3421 gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca 3481 tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac 3541 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga 3601 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga 3661 tctcatgctg gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac 3721 aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt 3781 tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc 3841 accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc 3901 aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc 3961 ctcaggttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat 4021 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt 4081 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct 4141 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc 4201 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc 4261 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc 4321 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc 4381 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg 4441 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata 4501 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta 4561 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc 4621 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg 4681 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt 4741 cctggccttt tgctggcctt ttgctcacat gttcttt //