LOCUS Exported 7252 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Flp-Dependent ChR2-EYFP. ACCESSION . VERSION . KEYWORDS pAAV-Ef1a-fDIO hChR2(H134R)-EYFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7252) AUTHORS Fenno LE, Mattis J, Ramakrishnan C, Hyun M, Lee SY, He M, Tucciarone J, Selimbeyoglu A, Berndt A, Grosenick L, Zalocusky KA, Bernstein H, Swanson H, Perry C, Diester I, Boyce FM, Bass CE, Neve R, Huang ZJ, Deisseroth K TITLE Targeting cells with single vectors using multiple-feature Boolean logic. JOURNAL Nat Methods. 2014 Jul;11(7):763-72. doi: 10.1038/nmeth.2996. Epub 2014 Jun 8. PUBMED 24908100 REFERENCE 2 (bases 1 to 7252) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7252 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 227..1405 /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 458..1396 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind 1353..1373 /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" protein_bind 1436..1469 /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" CDS complement(1565..2284) /codon_start=1 /product="enhanced YFP" /label=EYFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(1610..1631) /label=EGFP-C /note="EGFP, forward primer" primer_bind 1959..1978 /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2218..2239 /label=EGFP-N /note="EGFP, reverse primer" regulatory 2281..2290 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS complement(2300..3226) /codon_start=1 /product="H134R mutant of an N-terminal fragment of channelrhodopsin-2 from the alga Chlamydomonas reinhardtii (Nagel et al., 2005)" /label=ChR2(H134R) /note="blue light-activated cation channel" /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRGT NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEFKN PSMLYLATGHRVQWLRYAEWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI RKTTKLNIGGTEIEVETLVEDEAEAGAV" regulatory 3223..3232 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" protein_bind complement(3237..3270) /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" misc_feature 3379..3967 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(3432..3452) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3850..3861) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3999..4475 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(4198..4217) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 4515..4655 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 4515..4644 /label=AAV2 ITR rep_origin 4730..5185 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4817..4836) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 5027..5048 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(5202..5221) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 5321..5343 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(5381..5399) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 5467..5571 /gene="bla" /label=AmpR promoter CDS 5572..6432 /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(5790..5809) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 6603..7191 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 7092..7111 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt 61 ggtcgcccgg cctcagtgag cgagcgcgca gagagggagt ggccaactcc atcactaggg 121 gttcctgcgg ccgcacgcgt aagctttgca aagatggata aagttttaaa cagagaggaa 181 tctttgcagc taatggacct tctaggtctt gaaaggagtg ggaattggct ccggtgcccg 241 tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa 301 ttgaaccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg 361 gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa 421 cgttcttttt cgcaacgggt ttgccgccag aacacaggta agtgccgtgt gtggttcccg 481 cgggcctggc ctctttacgg gttatggccc ttgcgtgcct tgaattactt ccacctggct 541 gcagtacgtg attcttgatc ccgagcttcg ggttggaagt gggtgggaga gttcgaggcc 601 ttgcgcttaa ggagcccctt cgcctcgtgc ttgagttgag gcctggcctg ggcgctgggg 661 ccgccgcgtg cgaatctggt ggcaccttcg cgcctgtctc gctgctttcg ataagtctct 721 agccatttaa aatttttgat gacctgctgc gacgcttttt ttctggcaag atagtcttgt 781 aaatgcgggc caagatctgc acactggtat ttcggttttt ggggccgcgg gcggcgacgg 841 ggcccgtgcg tcccagcgca catgttcggc gaggcggggc ctgcgagcgc ggccaccgag 901 aatcggacgg gggtagtctc aagctggccg gcctgctctg gtgcctggcc tcgcgccgcc 961 gtgtatcgcc ccgccctggg cggcaaggct ggcccggtcg gcaccagttg cgtgagcgga 1021 aagatggccg cttcccggcc ctgctgcagg gagctcaaaa tggaggacgc ggcgctcggg 1081 agagcgggcg ggtgagtcac ccacacaaag gaaaagggcc tttccgtcct cagccgtcgc 1141 ttcatgtgac tccacggagt accgggcgcc gtccaggcac ctcgattagt tctcgagctt 1201 ttggagtacg tcgtctttag gttgggggga ggggttttat gcgatggagt ttccccacac 1261 tgagtgggtg gagactgaag ttaggccagc ttggcacttg atgtaattct ccttggaatt 1321 tgcccttttt gagtttggat cttggttcat tctcaagcct cagacagtgg ttcaaagttt 1381 ttttcttcca tttcaggtgt cgtgaggtac cggatcctct agagtcgact ccggagaagt 1441 tcctattctc tagaaagtat aggaacttcg cagaatggta gctggattgt agctgctatt 1501 agcaatatga aacctcttag aagttcctat tcttcaaaag gtataggaac ttcggcgcgc 1561 cctattactt gtacagctcg tccatgccga gagtgatccc ggcggcggtc acgaactcca 1621 gcaggaccat gtgatcgcgc ttctcgttgg ggtctttgct cagggcggac tggtagctca 1681 ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat gggggtgttc tgctggtagt 1741 ggtcggcgag ctgcacgctg ccgtcctcga tgttgtggcg gatcttgaag ttcaccttga 1801 tgccgttctt ctgcttgtcg gccatgatat agacgttgtg gctgttgtag ttgtactcca 1861 gcttgtgccc caggatgttg ccgtcctcct tgaagtcgat gcccttcagc tcgatgcggt 1921 tcaccagggt gtcgccctcg aacttcacct cggcgcgggt cttgtagttg ccgtcgtcct 1981 tgaagaagat ggtgcgctcc tggacgtagc cttcgggcat ggcggacttg aagaagtcgt 2041 gctgcttcat gtggtcgggg tagcgggcga agcactgcag gccgtagccg aaggtggtca 2101 cgagggtggg ccagggcacg ggcagcttgc cggtggtgca gatgaacttc agggtcagct 2161 tgccgtaggt ggcatcgccc tcgccctcgc cggacacgct gaacttgtgg ccgtttacgt 2221 cgccgtccag ctcgaccagg atgggcacca ccccggtgaa cagctcctcg cccttgctca 2281 ccatggtggc ggccgctggc acggctccgg cctcggcttc gtcttcgacg agagtctcga 2341 cctcgatctc cgttccgccg atgttcagtt tggtggtttt gcggatatct ccgtgaatca 2401 atatgtgctc gtggatcagg acgcgcaggt agtgtcccaa caacccccaa caatttttac 2461 tcatcagatc aataatcgtg tgacctacgg tggagccata gacgctcagg acgccaaaac 2521 cttcgggccc caaaatgaag agaattggga acatacccca gctcacgaaa aacagccatg 2581 ccatgccggt cacgacctgg cggcaccgac cctttggcac agtatgataa ccctcgatat 2641 atgctttggc ggcgtgaaaa aatgtgttcg cgccatagca caatccaaga caaaagaaga 2701 tgactttaac atagccggtt gccatggcgc tggtagcccc ccacacgata gtcccgatgt 2761 ctgagacaag gagtcccatg gttctcctgc tgtagtcgtt gctcaggccg gtgaggttgc 2821 tcaggcggat aaggatgaca ggacaagtga gcagccactc tgcatagcgc agccactgca 2881 cccggtgtcc tgtggcaagg tagagcatag agggattctt aaactcaaaa aagaactcga 2941 gaatcacctt aaccatttca atggcgcaca catagatctc ctcccagccg catgtagatt 3001 tccaggtttg gtaggcatag aacatcagca gcaaaatgct gaatcctgct gcaagccact 3061 gcaggacatt tgacgcggtc tgagcgccgt tcgtgccgcg agattcaatc catccggcac 3121 agtaacattg atcctcaggg accaggacgg acccgttcac caccacagga ttagtaacga 3181 acaaaagttc gcgtccgacg gcagacaaag cgccgccata gtccatggtg gctagcgaag 3241 ttcctatact ttctagagaa taggaacttc ttgccttaac ccagaaatta tcactgttat 3301 tctttagaat ggtgcaaaga gaagttccta taccttttga agaataggaa cttcgaattc 3361 gatatcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt 3421 attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat 3481 catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg 3541 tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt 3601 gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact 3661 ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc 3721 tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg 3781 tcctttcctt ggctgctcgc ctatgttgcc acctggattc tgcgcgggac gtccttctgc 3841 tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg 3901 cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc 3961 tccccgcatc gataccgagc gctgctcgag agatctacgg gtggcatccc tgtgacccct 4021 ccccagtgcc tctcctggcc ctggaagttg ccactccagt gcccaccagc cttgtcctaa 4081 taaaattaag ttgcatcatt ttgtctgact aggtgtcctt ctataatatt atggggtgga 4141 ggggggtggt atggagcaag gggcaagttg ggaagacaac ctgtagggcc tgcggggtct 4201 attgggaacc aagctggagt gcagtggcac aatcttggct cactgcaatc tccgcctcct 4261 gggttcaagc gattctcctg cctcagcctc ccgagttgtt gggattccag gcatgcatga 4321 ccaggctcag ctaatttttg tttttttggt agagacgggg tttcaccata ttggccaggc 4381 tggtctccaa ctcctaatct caggtgatct acccaccttg gcctcccaaa ttgctgggat 4441 tacaggcgtg aaccactgct cccttccctg tccttctgat tttgtaggta accacgtgcg 4501 gaccgagcgg ccgcaggaac ccctagtgat ggagttggcc actccctctc tgcgcgctcg 4561 ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc 4621 ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc ctgatgcggt attttctcct 4681 tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta cgcgccctgt 4741 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc 4801 agcgccttag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc 4861 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg 4921 cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc atcgccctga 4981 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc 5041 caaactggaa caacactcaa ctctatctcg ggctattctt ttgatttata agggattttg 5101 ccgatttcgg tctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt 5161 aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg ctctgatgcc 5221 gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt 5281 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag 5341 aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt 5401 ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga 5461 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc 5521 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt 5581 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct 5641 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt 5701 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt 5761 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac 5821 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac 5881 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct 5941 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg 6001 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg 6061 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca 6121 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa 6181 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt 6241 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc 6301 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg 6361 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt 6421 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt 6481 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc 6541 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct 6601 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta 6661 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc 6721 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac 6781 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct 6841 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat 6901 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg 6961 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa 7021 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg 7081 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga 7141 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc 7201 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gt //