LOCUS Exported 6418 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION An AAV packaging vector that expresses Cre-dependent membrane-localized AcGFP under control of the EF1a promoter.. ACCESSION . VERSION . KEYWORDS pOTTC710 - pAAV EF1a DIO Mem-AcGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6418) AUTHORS Sharpe MJ, Marchant NJ, Whitaker LR, Richie CT, Zhang YJ, Campbell EJ, Koivula PP, Necarsulmer JC, Mejias-Aponte C, Morales M, Pickel J, Smith JC, Niv Y, Shaham Y, Harvey BK, Schoenbaum G TITLE Lateral Hypothalamic GABAergic Neurons Encode Reward Predictions that Are Relayed to the Ventral Tegmental Area to Regulate Learning. JOURNAL Curr Biol. 2017 Jul 24;27(14):2089-2100.e5. doi: 10.1016/j.cub.2017.06.024. Epub 2017 Jul 6. PUBMED 28690111 REFERENCE 2 (bases 1 to 6418) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6418 /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" protein_bind 1450..1483 /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 1534..1567 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." CDS complement(1607..2320) /codon_start=1 /product="Aequorea coerulescens GFP" /label=AcGFP1 /note="mammalian codon-optimized" /translation="SKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLTLKF ICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFEDDGN YKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKNGIKVN FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMIYFGF VTAAAITHGMDELYK" protein_bind complement(2403..2436) /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(2487..2520) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." misc_feature 2545..3133 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2598..2618) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3016..3027) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3165..3641 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(3364..3383) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 3681..3821 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 3681..3810 /label=AAV2 ITR rep_origin 3896..4351 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3983..4002) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4193..4214 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4368..4387) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 4487..4509 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(4547..4565) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 4633..4737 /gene="bla" /label=AmpR promoter CDS 4738..5598 /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(4956..4975) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5769..6357 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6258..6277 /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 ctctagagtc 1441 gactccggaa taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga 1501 ttgtagctgc tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg 1561 aagttatggc gcgcctggct gattatgatc tagagtcgcg gccgcttcac ttgtacagct 1621 catccatgcc gtgggtgatg gcggcggcgg tcacgaagcc gaagtagatc atgtgatcgc 1681 gcttctcgtt ggggtccttg gacagggcgc tctgggtgga caggtagtgg ttatcgggca 1741 gcagcacagg gccatcgccg atgggggtat tctgctggta gtggtcggcc agctgcacgc 1801 tgccatcctc gatgttgtgg cggatcttga agttcacctt gatgccattc ttggccttgt 1861 cggtcatgat gtacacattg tgggcgttgt agttgtactc catcttattg cccaggatgt 1921 tgccatcctc cttgaaatcg gtgccggtca gctcgatgcg attcaccagg gtatcgccct 1981 cgaacttcac ctcggcgcgc gacttgtagt tgccgtcatc ctcgaagaag atggtgcgct 2041 cctggatgta gccctcaggc atggcgctct tgaagaagtc gtgctgcttc atgtgatcgg 2101 ggtagcgtga gaagcactgc acgccgtagc tcagggtggt caccagggtg ggccagggca 2161 caggcagctt gccggtggtg cagatgaact tcagggtcag cttgccgtag gtggcatcgc 2221 cctcgccctc gccgctcacg ctgaacttgt ggccattcac atcgccattc agctcgatca 2281 ggatgggcac gatgccggtg aacagctcgg cgcccttgct gatcttttgg tcctcatcat 2341 tcttttcaac ctgtttggtt cttctcatac agcacagcat ggtggcgacc ggtgggctag 2401 cgataacttc gtataaagta tcctatacga agttatttgc cttaacccag aaattatcac 2461 tgttattctt tagaatggtg caaagaataa cttcgtataa tgtatgctat acgaagttat 2521 gaattcgata tcaagcttat cgataatcaa cctctggatt acaaaatttg tgaaagattg 2581 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct 2641 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg 2701 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact 2761 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc 2821 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc 2881 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa 2941 tcatcgtcct ttccttggct gctcgcctat gttgccacct ggattctgcg cgggacgtcc 3001 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg 3061 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg 3121 gccgcctccc cgcatcgata ccgagcgctg ctcgagagat ctacgggtgg catccctgtg 3181 acccctcccc agtgcctctc ctggccctgg aagttgccac tccagtgccc accagccttg 3241 tcctaataaa attaagttgc atcattttgt ctgactaggt gtccttctat aatattatgg 3301 ggtggagggg ggtggtatgg agcaaggggc aagttgggaa gacaacctgt agggcctgcg 3361 gggtctattg ggaaccaagc tggagtgcag tggcacaatc ttggctcact gcaatctccg 3421 cctcctgggt tcaagcgatt ctcctgcctc agcctcccga gttgttggga ttccaggcat 3481 gcatgaccag gctcagctaa tttttgtttt tttggtagag acggggtttc accatattgg 3541 ccaggctggt ctccaactcc taatctcagg tgatctaccc accttggcct cccaaattgc 3601 tgggattaca ggcgtgaacc actgctccct tccctgtcct tctgattttg taggtaacca 3661 cgtgcggacc gagcggccgc aggaacccct agtgatggag ttggccactc cctctctgcg 3721 cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg 3781 ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga tgcggtattt 3841 tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc atagtacgcg 3901 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca 3961 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc 4021 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct 4081 ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg 4141 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc 4201 ttgttccaaa ctggaacaac actcaaccct atctcgggct attcttttga tttataaggg 4261 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg 4321 aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac aatctgctct 4381 gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg 4441 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg 4501 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc 4561 ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt 4621 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat 4681 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg 4741 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt 4801 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga 4861 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa 4921 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt 4981 attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt 5041 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc 5101 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga 5161 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat 5221 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct 5281 gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc 5341 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg 5401 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc 5461 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg 5521 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca 5581 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta 5641 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc 5701 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa 5761 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca 5821 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta 5881 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc 5941 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca 6001 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta 6061 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag 6121 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt 6181 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc 6241 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac 6301 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac 6361 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgt //