LOCUS Exported 7084 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION 2P-activatable bPAC for AAV. ACCESSION . VERSION . KEYWORDS pAAV_2pabPAC SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7084) AUTHORS Kinjo T, Watabe T, Kobachi K, Terai K, Matsuda M TITLE Single-Cell Activation of the cAMP-Signaling Pathway in 3D Tissues with FRET-Assisted Two-Photon Activation of bPAC. JOURNAL ACS Chem Biol. 2020 Oct 19. doi: 10.1021/acschembio.0c00333. PUBMED 33074647 REFERENCE 2 (bases 1 to 7084) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7084 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 201..504 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 505..708 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 659..679 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 705..729 /label=LNCX /note="Human CMV promoter, forward primer" CDS 3044..3100 /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 3101..3811 /codon_start=1 /product="monomeric teal (cyan) fluorescent protein (Ai et al., 2006)" /label=mTFP1 /note="mammalian codon-optimized" /translation="MVSKGEETTMGVIKPDMKIKLKMEGNVNGHAFVIEGEGEGKPYDG TNTINLEVKEGAPLPFSYDILTTAFAYGNRAFTKYPDDIPNYFKQSFPEGYSWERTMTF EDKGIVKVKSDISMEEDSFIYEIHLKGENFPPNGPVMQKKTTGWDASTERMYVRDGVLK GDVKHKLLLEGGGHHRVDFKTIYRAKKAVKLPDYHFVDHRIEILNHDKDYNKVTVYESA VARNSTDGMDELYK" polyA_signal 3826..4302 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(4025..4044) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 4340..4469 /label=AAV2 ITR (alternate) /note="Functional equivalent of wild-type AAV2 ITR" rep_origin complement(4531..5119) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4611..4630) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5290..6150) /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 5913..5932 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6151..6255) /gene="bla" /label=AmpR promoter primer_bind 6323..6341 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(6379..6401) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 6501..6520 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" rep_origin complement(6537..6992) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(6674..6695) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 6886..6905 /label=F1ori-R /note="F1 origin, reverse primer" repeat_region 7067..123 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 7078..123 /label=AAV2 ITR ORIGIN 1 ctcgctcgct cactgaggcc gcccgggcaa agcccgggcg tcgggcgacc tttggtcgcc 61 cggcctcagt gagcgagcga gcgcgcagag agggagtggc caactccatc actaggggtt 121 cctgcggcca agcgtggagc tagttattaa tagtaatcaa ttacggggtc attagttcat 181 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg 241 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata 301 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta 361 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc 421 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac 481 gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga 541 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg 601 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg 661 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctcg tttagtgaac 721 cgtcagatcg cctggagacg ccatccacgc tgttttgacc tccatagaag acaccgggac 781 cgatccagcc tccgcggatt cgaatcccgg ccgggaacgg tgcattggaa cgcggattcc 841 ccgtgccaag agtgacgtaa gtaccgccta tagagtctat aggcccacaa aaaatgcttt 901 cttcttttaa tatacttttt tgtttatctt atttctaata ctttccctaa tctctttctt 961 tcagggcaat aatgatacaa tgtatcatgc ctctttgcac cattctaaag aataacagtg 1021 ataatttctg ggttaaggca atagcaatat ttctgcatat aaatatttct gcatataaat 1081 tgtaactgat gtaagaggtt tcatattgct aatagcagct acaatccagc taccattctg 1141 cttttatttt atggttggga taaggctgga ttattctgag tccaagctag gcccttttgc 1201 taatcatgtt catacctctt atcttcctcc cacagctcct gggcaacgtg ctggtctgtg 1261 tgctggccca tcactttggc aaagaattgg gattcgaaca tcgattgaat tcaccatgat 1321 ggtgagcaag ggcgaggaga ccacaatggg cgtaatcaag cccgacatga agatcaagct 1381 gaagatggag ggcaacgtga atggccacgc cttcgtgatc gagggcgagg gcgagggcaa 1441 gccctacgac ggcaccaaca ccatcaacct ggaggtgaag gagggagccc ccctgccctt 1501 ctcctacgac attctgacca ccgcgttcgc ctacggcaac agggccttca ccaagtaccc 1561 cgacgacatc cccaactact tcaagcagtc cttccccgag ggctactctt gggagcgcac 1621 catgaccttc gaggacaagg gcatcgtgaa ggtgaagtcc gacatctcca tggaggagga 1681 ctccttcatc tacgagatac acctcaaggg cgagaacttc ccccccaacg gccccgtgat 1741 gcagaagaag accaccggct gggacgcctc caccgagagg atgtacgtgc gcgacggcgt 1801 gctgaagggc gacgtcaagc acaagctgct gctggagggc ggcggccacc accgcgttga 1861 cttcaagacc atctacaggg ccaagaaggc ggtgaagctg cccgactatc actttgtgga 1921 ccaccgcatc gagatcctga accacgacaa ggactacaac aaggtgaccg tttacgagag 1981 cgccgtggcc cgcatgatga agcggctggt gtacatcagc aagatcagcg gccacctgag 2041 cctggaagag atccagcgga tcggcaaggt gtccatcaag aacaaccagc gggacaacat 2101 caccggcgtg ctgctgtacc tgcagggcct gttcttccag atcctggaag gcgagaacga 2161 gaaggtggac aagctgtaca agaaaatcct ggtggacgac cggcacacca acatcctgtg 2221 cctgaaaacc gagtacgaca tcaccgaccg gatgttcccc aactgggcca tgaaaaccat 2281 caacctgaac gagaacagcg agctgatgat ccagcccatc aagagcctgc tgcagaccat 2341 cacccagagc caccgggtgc tggaaaagta catgcccgcc agagtgatct acctgatcaa 2401 ccagggcatc aaccccctga ccgtggagcc ccagctggtc gagaagatca tcttcttcag 2461 cgacatcctg gccttcagca ccctgaccga gaagctgccc gtgaacgagg tggtcatcct 2521 ggtcaaccgg tacttcagca tctgcacccg gatcatcagc gcctacggcg gcgaagtgac 2581 caagttcatc ggcgactgcg tgatggccag cttcaccaaa gagcagggcg acgccgccat 2641 ccggaccagc ctggacatca tctccgagct gaagcagctg cggcaccacg tggaggccac 2701 caaccccctg cacctgctgt acaccggcat cggcctgagc tacggccacg tgatcgaggg 2761 caacatgggc agcagcctga agatggacca caccctgctg ggggacgccg tgaacgtggc 2821 cgccaggctg gaagccctga caagacagct gccctacgcc ctggccttta cagccggcgt 2881 gaagaagtgc tgccaggccc agtggacctt catcaacctg ggagcccacc aggtcaaggg 2941 caagcaggaa gccatcgagg tctacaccgt gaatgaggcc cagaagtact acgacaccct 3001 gcagatcacc cagctgatcc ggcagaccct ggaaaacgac aaggcgacga attttagtct 3061 actgaaacaa gcgggagacg tggaggaaaa ccctggacct atggtgagca agggcgagga 3121 gaccacaatg ggcgtaatca agcccgacat gaagatcaag ctgaagatgg agggcaacgt 3181 gaatggccac gccttcgtga tcgagggcga gggcgagggc aagccctacg acggcaccaa 3241 caccatcaac ctggaggtga aggagggagc ccccctgccc ttctcctacg acattctgac 3301 caccgcgttc gcctacggca acagggcctt caccaagtac cccgacgaca tccccaacta 3361 cttcaagcag tccttccccg agggctactc ttgggagcgc accatgacct tcgaggacaa 3421 gggcatcgtg aaggtgaagt ccgacatctc catggaggag gactccttca tctacgagat 3481 acacctcaag ggcgagaact tcccccccaa cggccccgtg atgcagaaga agaccaccgg 3541 ctgggacgcc tccaccgaga ggatgtacgt gcgcgacggc gtgctgaagg gcgacgtcaa 3601 gcacaagctg ctgctggagg gcggcggcca ccaccgcgtt gacttcaaga ccatctacag 3661 ggccaagaag gcggtgaagc tgcccgacta tcactttgtg gaccaccgca tcgagatcct 3721 gaaccacgac aaggactaca acaaggtgac cgtttacgag agcgccgtgg cccgcaactc 3781 caccgacggc atggacgagc tgtacaagta actcgagaga tctacgggtg gcatccctgt 3841 gacccctccc cagtgcctct cctggccctg gaagttgcca ctccagtgcc caccagcctt 3901 gtcctaataa aattaagttg catcattttg tctgactagg tgtccttcta taatattatg 3961 gggtggaggg gggtggtatg gagcaagggg caagttggga agacaacctg tagggcctgc 4021 ggggtctatt gggaaccaag ctggagtgca gtggcacaat cttggctcac tgcaatctcc 4081 gcctcctggg ttcaagcgat tctcctgcct cagcctcccg agttgttggg attccaggca 4141 tgcatgacca ggctcagcta atttttgttt ttttggtaga gacggggttt caccatattg 4201 gccaggctgg tctccaactc ctaatctcag gtgatctacc caccttggcc tcccaaattg 4261 ctgggattac aggcgtgaac cactgctccc ttccctgtcc ttctgatttt gtaggtaacc 4321 acgtgcggac cgaggccgca ggaaccccta gtgatggagt tggccactcc ctctctgcgc 4381 gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg cggcctcagt 4441 gagcgagcga gcgcgcagct gcctgcagga catgtgagca aaaggccagc aaaaggccag 4501 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca 4561 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca 4621 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg 4681 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag 4741 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt 4801 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca 4861 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg 4921 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt 4981 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc 5041 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg 5101 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg 5161 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta 5221 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg 5281 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg 5341 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc 5401 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc 5461 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc 5521 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag 5581 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat 5641 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg 5701 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt 5761 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag 5821 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg 5881 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt 5941 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct 6001 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac 6061 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat 6121 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat 6181 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca 6241 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat 6301 tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt 6361 cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct 6421 gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg 6481 tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc accataaaat 6541 tgtaaacgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt 6601 taaccaatag accgaaatcg gcaaaatccc ttataaatca aaagaatagc ccgagataga 6661 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt 6721 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat cacccaaatc 6781 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg 6841 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa 6901 aggagcgggc gctaaggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc 6961 cgccgcgctt aatgcgccgc tacagggcgc gtactatggt tgctttgacg tatgcggtgt 7021 gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgcccctg caggcagctg 7081 cgcg //