LOCUS Exported 7028 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Intersectional viral expression of iC++-EYFP in cells expressing Flp AND NOT Cre. ACCESSION . VERSION . KEYWORDS pAAV-nEF-Coff/Fon-iC++-EYFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7028) AUTHORS Fenno LE, Ramakrishnan C, Kim YS, Evans KE, Lo M, Vesuna S, Inoue M, Cheung KYM, Yuen E, Pichamoorthy N, Hong ASO, Deisseroth K TITLE Comprehensive Dual- and Triple-Feature Intersectional Single-Vector Delivery of Diverse Functional Payloads to Cells of Behaving Mammals. JOURNAL Neuron. 2020 Jun 20. pii: S0896-6273(20)30432-3. doi: 10.1016/j.neuron.2020.06.003. PUBMED 32574559 REFERENCE 2 (bases 1 to 7028) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7028 /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 188..399 /label=EF-1-alpha core promoter /note="core promoter for human elongation factor EF-1-alpha" LTR 412..680 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from human T-cell leukemia virus (HTLV) type 1" CDS complement(837..1088) /codon_start=1 /product="C-terminal fragment of mVenus for use in bimolecular fluorescence complementation (BiFC) (Kodama and Hu, 2010)" /label=VC155 /translation="DKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNH YLSYQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYK" primer_bind complement(879..900) /label=EGFP-C /note="EGFP, forward primer" primer_bind 1228..1247 /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" protein_bind 1469..1502 /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 1553..1586 /label=loxN /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GTATACCT). loxN sites are compatible with each other, but incompatible with loxP or lox2272 sites." primer_bind 1699..1720 /label=EGFP-N /note="EGFP, reverse primer" protein_bind complement(2352..2385) /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(2436..2469) /label=loxN /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GTATACCT). loxN sites are compatible with each other, but incompatible with loxP or lox2272 sites." regulatory 2997..3006 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" misc_feature 3155..3743 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(3208..3228) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3626..3637) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3775..4251 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(3974..3993) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 4291..4431 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 4291..4420 /label=AAV2 ITR rep_origin 4506..4961 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4593..4612) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4803..4824 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4978..4997) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 5097..5119 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(5157..5175) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 5243..5347 /gene="bla" /label=AmpR promoter CDS 5348..6208 /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(5566..5585) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 6379..6967 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6868..6887 /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 cgcgtaagga tctgcgatcg ctccggtgcc 181 cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc 241 aattgaacgg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac 301 tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg 361 aacgttcttt ttcgcaacgg gtttgccgcc agaacacagc tgaagcttcg aggggctcgc 421 atctctcctt cacgcgcccg ccgccctacc tgaggccgcc atccacgccg gttgagtcgc 481 gttctgccgc ctcccgcctg tggtgcctcc tgaactgcgt ccgccgtcta ggtaagttta 541 aagctcaggt cgagaccggg cctttgtccg gcgctccctt ggagcctacc tagactcagc 601 cggctctcca cgctttgcct gaccctgctt gctcaactct acgtctttgt ttcgttttct 661 gttctgcgcc gttacagatc caagctgtga ccggcgccta cggatccgaa gttcctattc 721 ttcaaatagt ataggaactt cgcagagtag tagctggatt gtagctgcta ttagcaatct 781 aaaacctctt agaagttcct attcttcaaa aggtatagga acttcggcgc gccttacttg 841 tacagctcgt ccatgccgag agtgatcccg gcggcggtca cgaactccag caggaccatg 901 tgatcgcgct tctcgttggg gtctttgctc agggcggact ggtagctcag gtagtggttg 961 tcgggcagca gcacggggcc gtcgccgatg ggggtgttct gctggtagtg gtcggcgagc 1021 tgcacgctgc cgtcctcgat gttgtggcgg atcttgaagt tcaccttgat gccgttcttc 1081 tgcttgtcgg ccatgatata gacgttgtgg ctgttgtagt tgtactccag cttgtgcccc 1141 aggatgttgc cgtcctcctt gaagtcgatg cccttcagct cgatgcggtt caccagggtg 1201 tcgccctcga acttcacctc ggcgcgggtc ttgtagttgc cgtcgtcctt gaagaagatg 1261 gtgcgctcct ggacgtagcc ttcgggcatg gcggacttga agaagtcgtg ctgcttcatg 1321 tggtcggggt agcgggcgaa gcactgcagg ccgtagccga aggtggtcac gagggtgggc 1381 cagggcacgg gcagctgtgg gacgacatga cttaaccagt taaccatgtt ctctgaacat 1441 gagaagggct ggccactctc cagtcgacat aacttcgtat aggatacttt atacgaagtt 1501 atgcagaatg gtagctggat tgtagctgct attagcaata tgaaacctct taataacttc 1561 gtatagtata ccttatacga agttatacta gtcctcctgt actcaccttg ccggtggtgc 1621 agatgaactt cagggtcagc ttgccgtagg tggcatcgcc ctcgccctcg ccggacacgc 1681 tgaacttgtg gccgtttacg tcgccgtcca gctcgaccag gatgggcacc accccggtga 1741 acagctcctc gcccttgctc accacgttga tgtcgatctg gtccaggggg atgtactcgc 1801 cctcgctggt gatcctgctc ttggcggccg cgaccgcgcc agcctcggcc tcgtcctcca 1861 ccagcgtctc gacctcaatc tcagtgccac caatgttcaa tttggtggtc ttgcgaatgt 1921 cgccgtggat gaggatatgg ctgtggatca gcacgcgcag gtagtggccg agcagacccc 1981 agcactgctt cgacatcagg tcaatgatgg tgtggccgac gttggagccg tacctgctca 2041 ggacgccgaa gccctcgggg ccgaggatga acaggatggg gaacataccc catgatacga 2101 agaagagcca agccatgcca gtcaccacct ggcgacaccg gcccttcggc acggtgtggt 2161 acgcctcaat gtagaccttg gctgcgttga agaatgtgta gatgccgtag cacaggccca 2221 tcaggaagaa aatgacacgg acgtatccct aaacacatga gaaatagagt cataagcaca 2281 tgggtcacat acaggagata tgtatataac attaatacaa ttttataaaa aaaagggggg 2341 gcacaggtac cataacttcg tataaagtat cctatacgaa gttatttgcc ttaacccaga 2401 aattatcact gttattcttt agaatggtgc aaagaataac ttcgtataag gtatactata 2461 cgaagttatg ctagcaaccc cgacacttac cttggacagc gcggccgtgg tgccccacac 2521 gatcgtgccg atatctgaca ccagcagacc catggtacgc ttgttatagt cgttcgccag 2581 acccgtaagg ttgctcaggt ggataagaat gacagggcag gtcagcagcc agctcgcgta 2641 acgaagccac ctggtcttgt tgccgttgga tgagtagatc accgcaggtt cgtcaaagct 2701 atggaaatac tcgatgatga acttgatcat ctggatcgtg gccacgtaaa tattctccca 2761 gccgcaagta gacttccagg tctggtagcc gtagaacatc aggcagagcg ctgaaagcgc 2821 aaaagaaatc cactgcagaa tgttggcagc caacttctcg gcatttgttc cgttggattt 2881 aagccaagcc aagcagaagc actggccgtt gttcgggatg caaataacag aaccattgtt 2941 ctcaagagtg tatgaggttt ggaagagtcc gacggcagac aaagcgccgc catagtccat 3001 ggtggcgagc tcgaagttcc tatactattt gaagaatagg aacttcttgc cttaacccag 3061 aaattatcac tgttattctt tagaatggtg caaagagaag ttcctatacc ttttgaagaa 3121 taggaacttc gaattcgata tcaagcttat cgataatcaa cctctggatt acaaaatttg 3181 tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc 3241 tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta 3301 taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt 3361 ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca 3421 gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc 3481 ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt 3541 gtcggggaaa tcatcgtcct ttccttggct gctcgcctat gttgccacct ggattctgcg 3601 cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg 3661 cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat 3721 ctccctttgg gccgcctccc cgcatcgata ccgagcgctg ctcgagagat ctacgggtgg 3781 catccctgtg acccctcccc agtgcctctc ctggccctgg aagttgccac tccagtgccc 3841 accagccttg tcctaataaa attaagttgc atcattttgt ctgactaggt gtccttctat 3901 aatattatgg ggtggagggg ggtggtatgg agcaaggggc aagttgggaa gacaacctgt 3961 agggcctgcg gggtctattg ggaaccaagc tggagtgcag tggcacaatc ttggctcact 4021 gcaatctccg cctcctgggt tcaagcgatt ctcctgcctc agcctcccga gttgttggga 4081 ttccaggcat gcatgaccag gctcagctaa tttttgtttt tttggtagag acggggtttc 4141 accatattgg ccaggctggt ctccaactcc taatctcagg tgatctaccc accttggcct 4201 cccaaattgc tgggattaca ggcgtgaacc actgctccct tccctgtcct tctgattttg 4261 taggtaacca cgtgcggacc gagcggccgc aggaacccct agtgatggag ttggccactc 4321 cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg 4381 gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga 4441 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc 4501 atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt 4561 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct 4621 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg 4681 atttagtgct ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag 4741 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa 4801 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcgggct attcttttga 4861 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa 4921 atttaacgcg aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac 4981 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc 5041 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg 5101 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct 5161 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg 5221 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc 5281 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag 5341 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg 5401 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt 5461 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt 5521 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt 5581 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa 5641 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag 5701 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac 5761 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac 5821 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac 5881 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac 5941 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact 6001 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg 6061 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt 6121 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat 6181 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta 6241 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa 6301 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga 6361 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac 6421 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt 6481 tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc 6541 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat 6601 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag 6661 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc 6721 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag 6781 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac 6841 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg 6901 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct 6961 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc 7021 tcacatgt //