LOCUS Exported 6706 bp ds-DNA linear SYN 16-JUL-2021 DEFINITION For AAV based expression of CheRiff in Cre-Positive cells. ACCESSION . VERSION . KEYWORDS SL016: AAV-CAG-FLEX-CheRiff-GFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6706) AUTHORS Farhi SL, Parot VJ, Grama A, Yamagata M, Abdelfattah AS, Adam Y, Lou S, Jun Kim J, Campbell RE, Cox DD, Cohen AE TITLE Wide-area all-optical neurophysiology in acute brain slices. JOURNAL J Neurosci. 2019 Apr 5. pii: JNEUROSCI.0168-19.2019. doi: 10.1523/JNEUROSCI.0168-19.2019. PUBMED 30952812 REFERENCE 2 (bases 1 to 6706) AUTHORS . TITLE Direct Submission JOURNAL Exported Jul 16, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6706 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin complement(8..463) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(145..166) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 357..376 /label=F1ori-R /note="F1 origin, reverse primer" repeat_region 535..664 /label=AAV2 ITR polyA_signal 826..947 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(863..882) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 917..936 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" misc_feature 990..1578 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" CDS 1096..1107 /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind 1505..1525 /label=WPRE-R /note="WPRE, reverse primer" protein_bind complement(1669..1702) /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)." regulatory 1796..1805 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1802..2749 /codon_start=1 /product="blue light-activated optogenetic actuator derived from Scherffelia dubia channelrhodopsin (Hochbaum et al., 2014)" /label=CheRiff /translation="MGGAPAPDAHSAPPGNDSAGGSEYHAPAGYQVNPPYHPVHGYEEQ CSSIYIYYGALWEQETARGFQWFAVFLSALFLAFYGWHAYKASVGWEEVYVCSVELIKV ILEIYFEFTSPAMLFLYGGNITPWLRYAEWLLTCPVILIHLSNITGLSEAYNKRTMALL VSDLGTICMGVTAALATGWVKWLFYCIGLVYGTQTFYNAGIIYVESYYIMPAGGCKKLV LAMTAVYYSSWLMFPGLFIFGPEGMHTLSVAGSTIGHTIADLLSKNIWGLLGHFLRIKI HEHIIMYGDIRRPVSSQFLGRKVDVLAFVTEEDKV" CDS 2819..3532 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITLGMDELYK" primer_bind complement(2861..2882) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(3122..3141) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 3469..3490 /label=EGFP-C /note="EGFP, forward primer" protein_bind 3578..3611 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." protein_bind 3674..3707 /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)." primer_bind complement(3742..3761) /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" primer_bind complement(3823..3845) /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" promoter complement(4124..4401) /label=chicken beta-actin promoter rep_origin complement(4904..5492) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4984..5003) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5663..6523) /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 6286..6305 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6524..6628) /gene="bla" /label=AmpR promoter ORIGIN 1 gccacctaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc 61 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag 121 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg 181 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca 241 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa 301 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg 361 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta 421 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccattcagg 481 ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctgcgc 541 gctcgctcgc tcactgaggc cgcccgggca aagcccgggc gtcgggcgac ctttggtcgc 601 ccggcctcag tgagcgagcg agcgcgcaga gagggagtgg ccaactccat cactaggggt 661 tccttgtagt taatgattaa cccgccatgc tacttatcta cgtagccatg ctctaggaag 721 atccttatcg attttaccac atttgtagag gttttacttg ctttaaaaaa cctcccacat 781 ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca 841 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt 901 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgctcg 961 aagcggccgc ccgggtcgac ggtatcgatg cggggaggcg gcccaaaggg agatccgact 1021 cgtctgaggg cgaaggcgaa gacgcggaag aggccgcaga gccggcagca ggccgcggga 1081 aggaaggtcc gctggattga gggccgaagg gacgtagcag aaggacgtcc cgcgcagaat 1141 ccaggtggca acacaggcga gcagccaagg aaaggacgat gatttccccg acaacaccac 1201 ggaattgtca gtgcccaaca gccgagcccc tgtccagcag cgggcaaggc aggcggcgat 1261 gagttccgcc gtggcaatag ggagggggaa agcgaaagtc ccggaaagga gctgacaggt 1321 ggtggcaatg ccccaaccag tgggggttgc gtcagcaaac acagtgcaca ccacgccacg 1381 ttgcctgaca acgggccaca actcctcata aagagacagc aaccaggatt tatacaagga 1441 ggagaaaatg aaagccatac gggaagcaat agcatgatac aaaggcatta aagcagcgta 1501 tccacatagc gtaaaaggag caacatagtt aagaatacca gtcaatcttt cacaaatttt 1561 gtaatccaga ggttgattat cgataagctt gatatcgaag ttcgatggtg agatctggac 1621 tagagggtcg atggtgatgc ttggatcaaa ataggaagaa tcccaatgat aacttcgtat 1681 aaagtatcct atacgaagtt atactagaat tcggtacaat caagctggtt ctccatttgc 1741 actcaggagc acgggcaggc gagtggcccc tagttctggg ggcagcgggg gatccgccac 1801 catgggcgga gctcctgctc cagacgctca cagcgcccca cctggaaacg attctgccgg 1861 aggcagtgag taccatgccc cagctggata tcaagtgaat ccaccctacc accccgtgca 1921 tgggtatgag gaacagtgca gctccatcta catctactat ggggccctgt gggagcagga 1981 aacagctagg ggcttccagt ggtttgccgt gttcctgtct gccctgtttc tggctttcta 2041 cggctggcac gcctataagg ccagcgtggg atgggaggaa gtgtacgtgt gctccgtgga 2101 gctgatcaaa gtgattctgg agatctattt cgagttcacc agtcctgcta tgctgttcct 2161 gtacggaggg aacattaccc catggctgag atatgccgaa tggctgctga catgtcccgt 2221 gatcctgatt catctgtcta acatcaccgg cctgagtgag gcatacaata agcggacaat 2281 ggctctgctg gtgtccgacc tgggaactat ttgcatggga gtgacagccg ctctggccac 2341 tgggtgggtg aagtggctgt tttactgtat cggcctggtg tatggaaccc agacattcta 2401 caacgctgga atcatctacg tggagtctta ctatatcatg cctgccggcg gctgtaagaa 2461 actggtgctg gccatgactg ccgtgtacta ttctagttgg ctgatgtttc ccggcctgtt 2521 catctttggg cctgaaggca tgcacaccct gagcgtggct gggtccacta ttggccatac 2581 catcgccgac ctgctgtcca agaatatttg gggactgctg gggcacttcc tgcggatcaa 2641 aattcacgag catatcatta tgtacggcga tatcaggaga ccagtgagct cccagtttct 2701 gggacgcaag gtggacgtgc tggccttcgt gacagaggaa gataaagtgg cggccgccaa 2761 gagcaggatc accagcgagg gcgagtacat ccccctggac cagatcgaca tcaacgtggt 2821 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga 2881 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa 2941 gctgaccctg aagttcattt gcaccaccgg caagctgccc gtgccctggc ccaccctcgt 3001 gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca 3061 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa 3121 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa 3181 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct 3241 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat 3301 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca 3361 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct 3421 gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct 3481 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtaagaatt 3541 cgatatcaag cttatcgaaa gtaaggggat ccctagtata acttcgtata gcatacatta 3601 tacgaagtta tgggtcgatg gtgatgcttg gcaattcggg tcgatggtga agcattggtc 3661 ttcctatttt gatataactt cgtataggat actttatacg aagttatggg gatccaattc 3721 tttgccaaaa tgatgagaca gcacaataac cagcacgttg cccaggagct gtaggaaaaa 3781 gaagaaggca tgaacatggt tagcagaggc tctagagccg ccggtcacac gccagaagcc 3841 gaaccccgcc ctgccccgtc ccccccgaag gcagccgtcc ccccgcggac agccccgagc 3901 cgctcccccc gcacaaaggg ccctcccgga gcccctcaag gctttcacgc agccacagaa 3961 aagaaacaag ccgtcattaa accaagcgct aattacagcc cggaggagaa gggccgtccc 4021 gcccgctcac ctgtgggagt aacgcggtca gtcagagccg gggcgggcgg cgcgaggcgg 4081 cggcggagcg gggcacgggg cgaaggcagc gcgcagcgac tcccgcccgc cgcgcgcttc 4141 gctttttata gggccgccgc cgccgccgcc tcgccataaa aggaaacttt cggagcgcgc 4201 cgctctgatt ggctgccgcc gcaccccccc gccccgcccc cccccccccc cccccccccc 4261 cccccccccc cggcgcgcgc cccccccccc cccccgcccc catcgctgca caaaataatt 4321 aaaaaataaa taaatacaaa attgggggtg gggagggggg ggagatgggg agagtgaagc 4381 agaacgtggg gctcacctcg accatggtaa tagcgatgac taatacgtag atgtactgcc 4441 aagtaggaaa gtcccataag gtcatgtact gggcataatg ccaggcgggc catttaccgt 4501 cattgacgtc aatagggggc gtacttggca tatgatacac ttgatgtact gccaagtggg 4561 cagtttaccg taaatactcc acccattgac gtcaatggaa agtccctatt ggcgttacta 4621 tgggaacata cgtcattatt gacgtcaatg gtactcttcc tagagcatgg ctacgtagat 4681 aagtagcatg gcgggttaat cattaactac aaggaacccc tagtgatgga gttggccact 4741 ccctctctgc gcgctcgctc gctcgctgag gccgagcggt atcagctcac tcaaaggcgg 4801 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc 4861 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc 4921 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac 4981 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc 5041 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata 5101 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc 5161 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca 5221 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag 5281 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta 5341 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg 5401 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc 5461 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt 5521 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa 5581 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat 5641 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga 5701 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac 5761 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg 5821 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg 5881 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt 5941 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct 6001 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat 6061 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta 6121 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca 6181 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat 6241 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac 6301 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa 6361 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt 6421 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg 6481 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat 6541 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt 6601 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctaaattgt 6661 aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatc //