LOCUS Exported 5302 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION AAV expression of membrane bound EGFP from GfaABC1D promoter. ACCESSION . VERSION . KEYWORDS pAAV.GfaABC1D.PI.Lck-GFP.SV40 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5302) AUTHORS Shigetomi E, Bushong EA, Haustein MD, Tong X, Jackson-Weaver O, Kracun S, Xu J, Sofroniew MV, Ellisman MH, Khakh BS TITLE Imaging calcium microdomains within entire astrocyte territories and endfeet with GCaMPs expressed using adeno-associated viruses. JOURNAL J Gen Physiol. 2013 Apr 15. PUBMED 23589582 REFERENCE 2 (bases 1 to 5302) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5302 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..130 /label=AAV2 ITR intron 981..1113 /label=chimeric intron /note="chimera between introns from human beta-globin and immunoglobulin heavy chain genes" primer_bind 1096..1115 /label=pMT2-F /note="Synthetic intron, forward primer" promoter 1158..1176 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" regulatory 1304..1313 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1310..2029 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 1310..2026 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(1355..1376) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(1616..1635) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 1963..1984 /label=EGFP-C /note="EGFP, forward primer" polyA_signal 2069..2190 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2080..2099) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2134..2153 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 2352..2481 /label=AAV2 ITR primer_bind complement(2500..2517) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(2500..2516) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2509..2531) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 2658..3113 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2745..2764) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2955..2976 /label=F1ori-F /note="F1 origin, forward primer" promoter 3139..3243 /gene="bla" /label=AmpR promoter CDS 3244..4104 /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 complement(3462..3481) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 4275..4863 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4764..4783 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5017..5034 /label=L4440 /note="L4440 vector, forward primer" protein_bind 5151..5172 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 5187..5217 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5225..5241 /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)." primer_bind 5230..5252 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 5249..5265 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 5249..5265 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" ORIGIN 1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt 61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact 121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct 181 aggaagatct aacatatcct ggtgtggagt aggggacgct gctctgacag aggctcgggg 241 gcctgagctg gctctgtgag ctggggagga ggcagacagc caggccttgt ctgcaagcag 301 acctggcagc attgggctgg ccgcccccca gggcctcctc ttcatgccca gtgaatgact 361 caccttggca cagacacaat gttcggggtg ggcacagtgc ctgcttcccg ccgcacccca 421 gcccccctca aatgccttcc gagaagccca ttgagcaggg ggcttgcatt gcaccccagc 481 ctgacagcct ggcatcttgg gataaaagca gcacagcccc ctaggggctg cccttgctgt 541 gtggcgccac cggcggtgga gaacaaggct ctattcagcc tgtgcccagg aaaggggatc 601 aggggatgcc caggcatgga cagtgggtgg caggggggga gaggagggct gtctgcttcc 661 cagaagtcca aggacacaaa tgggtgaggg gagagctctc cccatagctg ggctgcggcc 721 caaccccacc ccctcaggct atgccagggg gtgttgccag gggcacccgg gcatcgccag 781 tctagcccac tccttcataa agccctcgca tcccaggagc gagcagagcc agagcaggtt 841 ggagaggaga cgcatcacct ccgctgctcg caagctttat tgcggtagtt tatcacagtt 901 aaattgctaa cgcagtcagt gcttctgaca caacagtctc gaacttaagc tgcagaagtt 961 ggtcgtgagg cactgggcag gtaagtatca aggttacaag acaggtttaa ggagaccaat 1021 agaaactggg cttgtcgaga cagagaagac tcttgcgttt ctgataggca cctattggtc 1081 ttactgacat ccactttgcc tttctctcca caggtgtcca ctcccagttc aattacagct 1141 cttaaggcta gagtacttaa tacgactcac tataggctag cctcgagaat tcaccatggg 1201 ctgtggctgc agctcaaacc ctgaagatga ctggatggag aacattgacg tgtgtgagaa 1261 ctgccattat cccccgccga agcttcgaat tgatccaccg gtcgccacca tggtgagcaa 1321 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa 1381 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac 1441 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac 1501 cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt 1561 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga 1621 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat 1681 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta 1741 caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt 1801 gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca 1861 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac 1921 ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt 1981 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtaat ctagagtcga 2041 cccgggcggc cgcttcgagc agacatgata agatacattg atgagtttgg acaaaccaca 2101 actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt 2161 gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca ttttatgttt 2221 caggttcagg gggagatgtg ggaggttttt taaagcaagt aaaacctcta caaatgtggt 2281 aaaatcgata aggatcttcc tagagcatgg ctacgtagat aagtagcatg gcgggttaat 2341 cattaactac aaggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc 2401 gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc 2461 agtgagcgag cgagcgcgca gccttaatta acctaattca ctggccgtcg ttttacaacg 2521 tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt 2581 cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag 2641 cctgaatggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt 2701 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt 2761 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc 2821 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga 2881 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc 2941 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt 3001 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct 3061 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc 3121 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat 3181 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag 3241 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt 3301 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt 3361 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc 3421 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta 3481 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac 3541 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa 3601 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg 3661 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc 3721 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg 3781 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta 3841 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg 3901 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg 3961 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc 4021 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt 4081 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt 4141 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc 4201 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag 4261 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa 4321 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg 4381 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag 4441 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg 4501 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga 4561 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc 4621 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc 4681 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga 4741 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt 4801 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg 4861 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac 4921 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga 4981 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg 5041 gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc 5101 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt 5161 tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt 5221 ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaga 5281 tttaattaag gccttaatta gg //