LOCUS Exported 8565 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pAAV-EF1a-FLEX-GTB SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8565) AUTHORS Haubensak W, Kunwar PS, Cai H, Ciocchi S, Wall NR, Ponnusamy R, Biag J, Dong HW, Deisseroth K, Callaway EM, Fanselow MS, Luthi A, Anderson DJ TITLE Genetic dissection of an amygdala microcircuit that gates conditioned fear. JOURNAL Nature. 2010 Nov 11. 468(7321):270-6. PUBMED 21068836 REFERENCE 2 (bases 1 to 8565) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8565 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..141 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 12..141 /label=AAV2 ITR promoter 242..1419 /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 473..1410 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind 1367..1387 /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" regulatory 1441..1450 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" protein_bind complement(1453..1486) /label=lox2272 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GGATACTT). lox2272 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." protein_bind complement(1549..1582) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." CDS complement(3173..3232) /codon_start=1 /product="2A peptide from equine rhinitis A virus polyprotein" /label=E2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="QCTNYALLKLAGDVESNPGP" CDS complement(3722..3775) /codon_start=1 /product="2A peptide from Thosea asigna virus capsid protein" /label=T2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="EGRGSLLTCGDVEENPGP" CDS complement(3791..4504) /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITLGMDELYK" CDS complement(3791..4504) /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITLGMDELYK" primer_bind complement(3833..3854) /label=EGFP-C /note="EGFP, forward primer" primer_bind 4182..4201 /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 4441..4462 /label=EGFP-N /note="EGFP, reverse primer" protein_bind 4513..4546 /label=lox2272 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GGATACTT). lox2272 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." protein_bind 4617..4650 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." misc_feature 4692..5280 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(4745..4765) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(5163..5174) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 5312..5788 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(5511..5530) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 5828..5968 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 5828..5957 /label=AAV2 ITR rep_origin 6043..6498 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(6130..6149) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 6340..6361 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(6515..6534) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 6634..6656 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(6694..6712) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 6780..6884 /gene="bla" /label=AmpR promoter CDS 6885..7745 /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(7103..7122) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 7916..8504 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 8405..8424 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc 61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca 121 actccatcac taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt 181 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat 241 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg 301 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag 361 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc 421 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc 481 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat 541 tacttccact ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg 601 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg 661 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct 721 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg 781 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc 841 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga 901 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct 961 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca 1021 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg 1081 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg 1141 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat 1201 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg 1261 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa 1321 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca 1381 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagcc 1441 gccaccatgg ccataacttc gtataaagta tcctatacga agttatatca aaataggaag 1501 accaatgctt caccatcgac ccgaattgcc aagcatcacc atcgacccat aacttcgtat 1561 aatgtatgct atacgaagtt atacttcaac tagttcacag tctggtctcg ccgccgctct 1621 tgtggctctc ccagctgctg atgatcttgc cgctctgggg ggtcacgctc acctctctgc 1681 cggtgcctct caggttgtgc tgggtgggct cgcttctgtt cactcttctg cagcaggtca 1741 tcaggaagat gatcagcatc agggcggtca gggcgccggc gctcagcagc acgtacttgc 1801 cccagttggg caggcccagg tccacgccgc tcacctggtt gtgcacgtcg ggcaggtgca 1861 cctccacgaa gtcctcggcc tcgtcgccgt ccttgaacac ggtgctgggg tcggccaggg 1921 ggtgcaccag ggggatcacg ctgctctcca gcagctccat gtgctgctgc agcaggctgc 1981 tctgcatctc ggggatcagc acgttgccgt cggggcccag gatgatgccg ttgaagaaca 2041 cgccgttcac gtgggggtgg catctgccgc ccactctcag gcagcccttg ctgggcagga 2101 tctcgttcca ggttctcacg ctcttgtagt gggcgtcggc ctccatcagg gtcttgttga 2161 agatggtgta ggccttgccg aagccgggca ccagctttct caggtggctc agtcttctga 2221 agctcacgct cttggtggtc atgatgctct ccagggcgtc caggcactcc tctctctttc 2281 tcaccagctc ctccaccacc aggtgctcga tctcgtcgct tctgaagtcg tgcaggttca 2341 ccagcttgtc gggggggcac cacttggtct cgttgctggt ctgcatgctc acccaggtgc 2401 cgtccatcag tctcaggccc agcacgccgc acagcttcag cttgcaggcg cccttcaggc 2461 tcttgtacag gcctctctcg tccacgaagc cgcaggtctc gctgcccttg ctggctctct 2521 tgcctctgct gttggtgaag atgtcgcagc tcatgcccag tctggggttc tcgggcatcc 2581 agatggtgta gtcgtggttg gtgctgcagt aggtgctgct cacggccacg ccgctgcact 2641 tgccgctggg gaacactctg ctgtgcaggc ttctgtcgta ggggtccagg tcggccacgc 2701 tggggctgat gatcaccagg ctctccttgg tggtcttcac ggttctcagc catctgtagt 2761 cggggtaggg gttgtgcagg ctctcctcgt atctggggtc gccggccatc ttccagttgt 2821 aggcggctct gcaggcgtcg ggggtgggtc tgaagtgctt tctcttgaag gtggtggtca 2881 cgtagcccac gaagttggtg taggtctcgg cctcggtcac cacgccggtg caggtgaagc 2941 cgttcacctt gatggccagg atgtagccca ccttcagctc catgtagctg aagccgctca 3001 ggttggtgca gccctcgtcc tccaccacca ggttgttggg gcagctcagg tggtggatgt 3061 cgatggggct ccaggggccc agcttgtcgg ggatggtgta gatggggaac ttgccgaagc 3121 acagggggaa caccagcagg ggcacgaaca gcagggcctg gggcactccg gaaggaccgg 3181 ggttactttc aacatcgcca gcgagtttca acaaagcgta gttagtacat tgaccactac 3241 cgctagcgga gaacaagtct gcctggctct tgtcaggcac cagcagctcc ttggatgcgc 3301 tttcaagact gaagatgtca gacctgcttt ttgctttgga cttcccccat gcagcgatac 3361 cacccacagc taccaggcag cacaggagca ctgcagtgat cagcatccac atgcggccgt 3421 gattcctggc tggcagagca cgtccaggag caggggcagt gggagcctct gtgccgttgt 3481 ccgtgggcac cgcggggatc gcgctggtcc cgcagcccca ctcgtcccgc ccgtcgtcgc 3541 agtcggggtg tccgtcgcac agccagtcct gcgggtaaca ctccccgtgg gcaccgggcg 3601 gctccgagca gcggaactga ccgggggggc aacgggacaa agaaccgtta ccggacccgt 3661 taccggtcac gttaccgggc agcagcagca gcagcagcgc gggcagcagc cgcgcatcga 3721 ttgggccagg attctcctcg acgtcaccgc atgttagcag acttcctctg ccctctccac 3781 taccaagctt cttgtacagc tcgtccatgc cgagagtgat cccggcggcg gtcacgaact 3841 ccagcaggac catgtgatcg cgcttctcgt tggggtcttt gctcagggcg gactgggtgc 3901 tcaggtagtg gttgtcgggc agcagcacgg ggccgtcgcc gatgggggtg ttctgctggt 3961 agtggtcggc gagctgcacg ctgccgtcct cgatgttgtg gcggatcttg aagttcacct 4021 tgatgccgtt cttctgcttg tcggccatga tatagacgtt gtggctgttg tagttgtact 4081 ccagcttgtg ccccaggatg ttgccgtcct ccttgaagtc gatgcccttc agctcgatgc 4141 ggttcaccag ggtgtcgccc tcgaacttca cctcggcgcg ggtcttgtag ttgccgtcgt 4201 ccttgaagaa gatggtgcgc tcctggacgt agccttcggg catggcggac ttgaagaagt 4261 cgtgctgctt catgtggtcg gggtagcggc tgaagcactg cacgccgtag gtcagggtgg 4321 tcacgagggt gggccagggc acgggcagct tgccggtggt gcagatgaac ttcagggtca 4381 gcttgccgta ggtggcatcg ccctcgccct cgccggacac gctgaacttg tggccgttta 4441 cgtcgccgtc cagctcgacc aggatgggca ccaccccggt gaacagctcc tcgcccttgc 4501 tcacctcgac gcataacttc gtataggata ctttatacga agttatcttt gggattcttc 4561 ctattttgat ccaagcatca ccatcgaccc tctagtccag atctcaccat cgacccataa 4621 cttcgtatag catacattat acgaagttat gtccctcgaa gaggttcgaa ttcgatatca 4681 agcttatcga taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta 4741 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta 4801 ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt 4861 atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg 4921 caacccccac tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt 4981 tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag 5041 gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc 5101 cttggctgct cgcctatgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc 5161 cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc 5221 ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc 5281 atcgataccg agcgctgctc gagagatcta cgggtggcat ccctgtgacc cctccccagt 5341 gcctctcctg gccctggaag ttgccactcc agtgcccacc agccttgtcc taataaaatt 5401 aagttgcatc attttgtctg actaggtgtc cttctataat attatggggt ggaggggggt 5461 ggtatggagc aaggggcaag ttgggaagac aacctgtagg gcctgcgggg tctattggga 5521 accaagctgg agtgcagtgg cacaatcttg gctcactgca atctccgcct cctgggttca 5581 agcgattctc ctgcctcagc ctcccgagtt gttgggattc caggcatgca tgaccaggct 5641 cagctaattt ttgttttttt ggtagagacg gggtttcacc atattggcca ggctggtctc 5701 caactcctaa tctcaggtga tctacccacc ttggcctccc aaattgctgg gattacaggc 5761 gtgaaccact gctcccttcc ctgtccttct gattttgtag gtaaccacgt gcggaccgag 5821 cggccgcagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc 5881 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg 5941 agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat 6001 ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg 6061 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc 6121 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc 6181 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg 6241 accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg 6301 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg 6361 gaacaacact caaccctatc tcgggctatt cttttgattt ataagggatt ttgccgattt 6421 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa 6481 tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt 6541 taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc 6601 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt 6661 caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg 6721 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc 6781 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac 6841 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt 6901 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag 6961 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg 7021 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa 7081 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc 7141 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag 7201 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa 7261 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc 7321 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg 7381 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa 7441 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa 7501 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg 7561 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag 7621 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg 7681 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt 7741 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt 7801 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac 7861 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag 7921 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg 7981 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca 8041 gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga 8101 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca 8161 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc 8221 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca 8281 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa 8341 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc 8401 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc 8461 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg 8521 cctttttacg gttcctggcc ttttgctggc cttttgctca catgt //