LOCUS Exported 7598 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION cGMP FRET biosensor. ACCESSION . VERSION . KEYWORDS pCX-ThPDE5VV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7598) AUTHORS Ros O, Zagar Y, Ribes S, Baudet S, Loulier K, Couvet S, Ladarre D, Aghaie A, Louail A, Petit C, Mechulam Y, Lenkei Z, Nicol X TITLE SponGee: A Genetic Tool for Subcellular and Cell-Specific cGMP Manipulation. JOURNAL Cell Rep. 2019 Jun 25;27(13):4003-4012.e6. doi: 10.1016/j.celrep.2019.05.102. PUBMED 31242429 REFERENCE 2 (bases 1 to 7598) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7598 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 4..383 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 385..660 /label=chicken beta-actin promoter intron 661..1677 /label=chimeric intron /note="chimera between introns from chicken beta-actin and rabbit beta-globin" primer_bind 1601..1623 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 1685..1704 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" CDS 2985..3719 /codon_start=1 /product="circularly permuted form of the Venus yellow fluorescent protein (Nagai et al., 2004)" /label=cp173Venus /note="efficient FRET acceptor for CFP" /translation="MDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRD HMVLLEFVTAAGITLGMDELYKGGSGGMVSKGEELFTGVVPILVELDGDVNGHKFSVSG EGEGDATYGKLTLKLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPE GYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHN VYITADKQKNGIKANFKIRHNIE" CDS 3777..4493 /codon_start=1 /product="yellow fluorescent protein (YFP) with fast and efficient maturation (Nagai et al., 2002)" /label=Venus /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(4619..4638) /label=Bglob-pA-R /note="Rabbit beta-globin polyA region, reverse primer" polyA_signal 4684..4739 /label=-beta-globin poly(A) signal /note="rabbit beta-globin polyadenylation signal (Gil and Proudfoot, 1987)" primer_bind complement(4738..4757) /label=rbglobpA-R /note="Rabbit beta-globin polyA, reverse primer. Also called rb-glob-pA-term-R" primer_bind complement(5100..5116) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(5100..5116) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(5113..5135) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 5124..5140 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." promoter complement(5148..5178) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5193..5214 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 5273..5468 /label=SV40 promoter /note="SV40 early promoter" rep_origin 5319..5454 /label=SV40 ori /note="SV40 origin of replication" primer_bind 5381..5400 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" polyA_signal 5474..5608 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5511..5530) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 5565..5584 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(5676..5693) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5847..6435) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5927..5946) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6606..7466) /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 7229..7248 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7467..7571) /gene="bla" /label=AmpR promoter ORIGIN 1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata 61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc 121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag 181 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac 241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg 301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg 361 tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca 421 tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag 481 cgatgggggc gggggggggg ggggcgcgcg ccaggcgggg cggggcgggg cgaggggcgg 541 ggcggggcga ggcggagagg tgcggcggca gccaatcaga gcggcgcgct ccgaaagttt 601 ccttttatgg cgaggcggcg gcggcggcgg ccctataaaa agcgaagcgc gcggcgggcg 661 ggagtcgctg cgcgctgcct tcgccccgtg ccccgctccg ccgccgcctc gcgccgcccg 721 ccccggctct gactgaccgc gttactccca caggtgagcg ggcgggacgg cccttctcct 781 ccgggctgta attagcgctt ggtttaatga cggcttgttt cttttctgtg gctgcgtgaa 841 agccttgagg ggctccggga gggccctttg tgcgggggga gcggctcggg gggtgcgtgc 901 gtgtgtgtgt gcgtggggag cgccgcgtgc ggctccgcgc tgcccggcgg ctgtgagcgc 961 tgcgggcgcg gcgcggggct ttgtgcgctc cgcagtgtgc gcgaggggag cgcggccggg 1021 ggcggtgccc cgcggtgcgg ggggggctgc gaggggaaca aaggctgcgt gcggggtgtg 1081 tgcgtggggg ggtgagcagg gggtgtgggc gcgtcggtcg ggctgcaacc ccccctgcac 1141 ccccctcccc gagttgctga gcacggcccg gcttcgggtg cggggctccg tacggggcgt 1201 ggcgcggggc tcgccgtgcc gggcgggggg tggcggcagg tgggggtgcc gggcggggcg 1261 gggccgcctc gggccgggga gggctcgggg gaggggcgcg gcggcccccg gagcgccggc 1321 ggctgtcgag gcgcggcgag ccgcagccat tgccttttat ggtaatcgtg cgagagggcg 1381 cagggacttc ctttgtccca aatctgtgcg gagccgaaat ctgggaggcg ccgccgcacc 1441 ccctctagcg ggcgcggggc gaagcggtgc ggcgccggca ggaaggaaat gggcggggag 1501 ggccttcgtg cgtcgccgcg ccgccgtccc cttctccctc tccagcctcg gggctgtccg 1561 cggggggacg gctgccttcg ggggggacgg ggcagggcgg ggttcggctt ctggcgtgtg 1621 accggcggct ctagagcctc tgctaaccat gttcatgcct tcttcttttt cctacagctc 1681 ctgggcaacg tgctggttat tgtgctgtct catcattttg gcaaagaatt ctgataaact 1741 taagcttggt accgagctcg gatccactag tccagtgtgg tggaattctg cagatatcca 1801 gcacagtggc ggccgcaatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca 1861 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg 1921 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc 1981 ccgtgccctg gcccaccctc gtgaccaccc tgtcctgggg cgtgcagtgc ttcgcccgct 2041 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc 2101 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt 2161 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg 2221 gcaacatcct ggggcacaag ctggagtaca actacatcag cgacaacgtc tatatcaccg 2281 ccgacaagca gaagaacggc atcaaggcca acttcaagat ccgccacaac atcgaggacg 2341 gcggcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc 2401 tgctgcccga caaccactac ctgagcaccc agtccaagct gagcaaagac cccaacgaga 2461 agcgcgatca catggtcctg ctggagttcg tgaccgccgc ccgatgggaa ttagtgaagg 2521 atatttctag tcatttggat gtcacagcct tatgtcacaa aattttcttg catatccatg 2581 gactgatatc tgctgaccgc tattccctgt tccttgtctg tgaagacagc tccaatgaca 2641 agtttcttat cagccgcctc tttgatgttg ctgaaggttc aacactggaa gaagtttcaa 2701 ataactgtat ccgcttagaa tggaacaaag gcattgtggg acatgtggca gcgcttggtg 2761 agcccttgaa catcaaagat gcatatgagg atcctcggtt caatgcagaa gttgaccaaa 2821 ttacaggcta caagacacaa agcattcttt gtatgccaat taagaatcat agggaagagg 2881 ttgttggtgt agcccaggcc atcaacaaga aatcaggaaa cggtgggaca tttactgaaa 2941 aagatgaaaa ggactttgct gcttatttgg cagctagcga gctcatggac ggcggcgtgc 3001 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg 3061 acaaccacta cctgagctac cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc 3121 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt 3181 acaagggtgg cagcggtggc atggtgagca agggcgagga gctgttcacc ggggtggtgc 3241 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg 3301 gcgagggcga tgccacctac ggcaagctga ccctgaagct gatctgcacc accggcaagc 3361 tgcccgtgcc ctggcccacc ctcgtgacca ccctgggcta cggcctgcag tgcttcgccc 3421 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg 3481 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga 3541 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg 3601 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca 3661 ccgccgacaa gcagaagaac ggcatcaagg ccaacttcaa gatccgccac aacatcgagc 3721 cgaattttgt cttcctgatc ggcgccgcag gaatactctt cgtatctagc gagctcatgg 3781 tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg 3841 acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca 3901 agctgaccct gaagctgatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg 3961 tgaccactct gggctacggc ctacagtgct tcgcccgcta ccccgaccac atgaagcagc 4021 acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca 4081 aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga 4141 accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc 4201 tggagtacaa ctacaacagc cacaacgtct atatcaccgc cgacaagcag aagaacggca 4261 tcaaggccaa cttcaagatc cgccacaaca tcgaggacgg cggcgtgcag ctcgccgacc 4321 actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc 4381 tgagctacca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc 4441 tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagccgaatt 4501 ccctcgaggt taacgcggcg cgcctctaat ctagcgaatt cactcctcag gtgcaggctg 4561 cctatcagaa ggtggtggct ggtgtggcca atgccctggc tcacaaatac cactgagatc 4621 tttttccctc tgccaaaaat tatggggaca tcatgaagcc ccttgagcat ctgacttctg 4681 gctaataaag gaaatttatt ttcattgcaa tagtgtgttg gaattttttg tgtctctcac 4741 tcggaaggac atatgggagg gcaaatcatt taaaacatca gaatgagtat ttggtttaga 4801 gtttggcaac atatgcccat atgctggctg ccatgaacaa aggttggcta taaagaggtc 4861 atcagtatat gaaacagccc cctgctgtcc attccttatt ccatagaaaa gccttgactt 4921 gaggttagat tttttttata ttttgttttg tgttattttt ttctttaaca tccctaaaat 4981 tttccttaca tgttttacta gccagatttt tcctcctctc ctgactactc ccagtcatag 5041 ctgtccctct tctcttatgg agatccctcg acctgcagcc caagcttggc gtaatcatgg 5101 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc 5161 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg 5221 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagcggat ccgcatctca 5281 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca 5341 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg 5401 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct 5461 tttgcaaaaa gctaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat 5521 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact 5581 catcaatgta tcttatcatg tctggatccg ctgcattaat gaatcggcca acgcgcgggg 5641 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg 5701 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca 5761 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac 5821 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac 5881 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg 5941 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac 6001 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat 6061 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag 6121 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac 6181 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt 6241 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt 6301 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc 6361 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga 6421 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac 6481 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc 6541 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct 6601 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca 6661 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct 6721 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca 6781 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc 6841 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg 6901 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct 6961 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa 7021 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta 7081 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc 7141 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg 7201 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa 7261 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg 7321 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc 7381 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg 7441 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat 7501 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata 7561 ggggttccgc gcacatttcc ccgaaaagtg ccacctgg //