LOCUS Exported 7683 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Cry1 transcription fluorescence reporter. ACCESSION . VERSION . KEYWORDS AAV-P(Cry1)-DIO-intron336-Venus-NLS-D2 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7683) AUTHORS Mei L, Fan Y, Lv X, Welsh DK, Zhan C, Zhang EE TITLE Long-term in vivo recording of circadian rhythms in brains of freely moving mice. JOURNAL Proc Natl Acad Sci U S A. 2018 Apr 2. pii: 1717735115. doi: 10.1073/pnas.1717735115. PUBMED 29610316 REFERENCE 2 (bases 1 to 7683) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7683 /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 protein_bind 1733..1766 /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 1817..1850 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." CDS complement(2000..2020) /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS complement(2024..2737) /codon_start=1 /product="Venus YFP with monomerizing A206K mutation (Nagai et al., 2002; Kremers et al., 2006)" /label=mVenus /note="mammalian codon-optimized" /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK LICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLE FVTAAGITLGMDELYK" primer_bind complement(2066..2087) /label=EGFP-C /note="EGFP, forward primer" primer_bind 2415..2434 /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2674..2695 /label=EGFP-N /note="EGFP, reverse primer" CDS complement(2747..2767) /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQG" CDS complement(2792..2836) /codon_start=1 /product="peptide tag that allows for enzymatic biotinylation" /label=AviTag(TM) /translation="GLNDIFEAQKIEWHE" CDS complement(2858..2923) /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" CDS complement(3314..3337) /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" regulatory 3337..3346 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" promoter complement(3384..3402) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(3385..3402) /label=SP6 /note="SP6 promoter, forward primer" promoter complement(3408..3426) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(3469..3488) /label=pMT2-F /note="Synthetic intron, forward primer" intron 3471..3603 /label=chimeric intron /note="chimera between introns from human beta-globin and immunoglobulin heavy chain genes" protein_bind complement(3668..3701) /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(3752..3785) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." misc_feature 3810..4398 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(3863..3883) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(4281..4292) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 4430..4906 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(4629..4648) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 4946..5086 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 4946..5075 /label=AAV2 ITR rep_origin 5161..5616 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(5248..5267) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 5458..5479 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(5633..5652) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 5752..5774 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(5812..5830) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 5898..6002 /gene="bla" /label=AmpR promoter CDS 6003..6863 /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(6221..6240) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 7034..7622 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 7523..7542 /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 cgcgtgtaaa gatgcacatg tggccctgtg 181 ttctcattag acttcaattg ctttgataaa acatcatgag caaaagcaac ttggccagga 241 aggaaggggt ttgttgccgt ttacagttaa taatccatcg tggggagaag tcagggcaga 301 aattcaaagc agaaacctgg agatagaggg ctgaagcaga agccatggag gaaagccgtt 361 ttggttccct gtggcttcct cagcctgctt ccatatacaa cccgcctagg gctaccagcc 421 taatggtggc gccacacaca ctggtcgggt ccttccacat cactcattaa tcaagaaagt 481 gcccagactt ccctacaggc aacctgatgg aagtctttac tacaattaag gttcctcttc 541 ccatttaact tcagttgaag aaaaaaaaaa aaaaaaaaaa aaaaagcaaa gcaaaaagcc 601 caaaaaccta atcaaaatac cttctaaact acacttcatg taatagatta gagttgataa 661 attgacgatg tatctacagt attatgaaat gctgaatatc tgagatcgga cctgtcttct 721 ccagagcttg ggcaaaggtg cccgccccaa acagtaaaaa aggctaacgg cgtgatttct 781 gctcctttca gagcagggga ctccgattaa cacctgattg aagctgggtg agaggcagga 841 ctgtgcctcg gttgcctctc acccagccca gcaataactc tcagggcaca cggaactggc 901 ttcagacctt ccacacaacc cataggaaga gccctgggcg cgctcccgcg tcctagcctc 961 ttctcttcca ccgcgagagc gcagcgcacg cgcgccgtgc ccggcagagc gcgcgcccgc 1021 gagctccccg cagccccgtg ctcatccggg tccccgcgag cctcggccaa tggaggcggc 1081 tcccggcccc gggagcagaa ctatgcctcc tcccccttct cgcggcctgt ggcccccgcg 1141 agcggcctgg ggctcaggag acccgcagag gctgcgagtc cggatcccac gcgagaactc 1201 aggtcgtgag cgcccagcga tcgcaaccgg tcccgaagca ccgggcgcgg cggcgcacgc 1261 gggggtctga gccagtgtag taaacacact tcagaaacgt gaggtgccgg tggtcacgtg 1321 gggtgcatgc cgcccaatga gaagccgggg gcggggcgga ggccgctgac gcggcggagt 1381 cacccgggca gcctcgggac cggtgaccgg caggcgagcg tcccgcggcc tggagctcta 1441 gcccgcgctc tccgagctcg ctgcccactc cgggacgcac aggccgcgtc gcgcggaggc 1501 cgcgtgagct cgtgtccgtt cgtggaggcc gcctcgggcg cgccaccgtc tcctcgcggg 1561 tcgcctcggt tgggcgaggt ttccgtgacg agcctcccgg ggctgctgcg accagcccgg 1621 gccgtcgtgg cggctcgggc cccgcgcgct cgcttgcgct ccgggggtct caggccctgg 1681 aaggtatgcg tgtcctccgg acgagatctg cgatctaagt gtcgactccg gaataacttc 1741 gtataggata ctttatacga agttatgcag aatggtagct ggattgtagc tgctattagc 1801 aatatgaaac ctcttaataa cttcgtatag catacattat acgaagttat ggctacacat 1861 tgatcctagc agaagcacag gctgcagggt gacggtccat cccgctctcc tgggcacaag 1921 acatgggcag cgtgccatca tcctgctcct ccacctccgg cgggaagcca tggctaagct 1981 tagaattcac ggggtcttct acctttctct tcttttttgg aggcttgtac agctcgtcca 2041 tgccgagagt gatcccggcg gcggtcacga actccagcag gaccatgtga tcgcgcttct 2101 cgttggggtc tttgctcagt ttggactggt agctcaggta gtggttgtcg ggcagcagca 2161 cggggccgtc gccgatgggg gtgttctgct ggtagtggtc ggcgagctgc acgccgccgt 2221 cctcgatgtt gtggcggatc ttgaagtttg ccttgatgcc gttcttctgc ttgtcggcgg 2281 tgatatagac gttgtggctg ttgtagttgt actccagctt gtgccccagg atgttgccgt 2341 cctccttgaa gtcgatgccc ttcagctcga tgcggttcac cagggtgtcg ccctcgaact 2401 tcacctcggc gcgggtcttg tagttgccgt cgtccttgaa gaagatggtg cgctcctgga 2461 cgtagccttc gggcatggcg gacttgaaga agtcgtgctg cttcatgtgg tcggggtagc 2521 gggcgaagca ctgcaggccg tagccgaggg tggtcacgag ggtgggccag ggcacgggca 2581 gcttgccggt ggtgcagatc aacttcaggg tcagcttgcc gtaggtggca tcgccctcgc 2641 cctcgccgga cacgctgaac ttgtggccgt ttacgtcgcc gtccagctcg accaggatgg 2701 gcaccacccc ggtgaacagc tcctcgccct tgctcacacc tgcagggccc tgaaaataca 2761 ggttttcgga tcctgaaccg cctgccgatc cctcatgcca ctctattttc tgagcttcga 2821 aaatatcgtt caggccgcta gctgcggccg ctccaccctt atcgtcatcg tccttgtagt 2881 cgatgtcatg atctttataa tcaccgtcat ggtctttgta gtccatggtg ggtacctctt 2941 gacagctcta ccttttacta ctataaaaac gtacatgtag taacttcctt atgccacttc 3001 caaaagatga tttcaacatg aatcaacaga aatagtttct aaagtaggtc atcgctatag 3061 caaccagtgg gtggtttagc atactcccat cacaatgacc tactttctag tctgtgctag 3121 gacagcttat ataactagta tttctagttc tgtctataaa atagtctgaa aaaggcaatg 3181 gcaagataca acataatccc actgctactg aaaacaagac attatgaatt catataacac 3241 agtcatctgt cctatgccat agtttctaca ctgtgtcata acaaccatct cgattaggga 3301 taacagggta attcttgtca tcgtcatcct tgtagtccat ggtggcctct ccttcgagtg 3361 caaaaagaac aagtagcttg tattctatag tgtcacctaa atgctagcct atagtgagtc 3421 gtattaagta ctctagcctt aagagctgta attgaactgg gagtggacac ctgtggagag 3481 aaaggcaaag tggatgtcag taagaccaat aggtgcctat cagaaacgca agagtcttct 3541 ctgtctcgac aagcccagtt tctattggtc tccttaaacc tgtcttgtaa ccttgatact 3601 tacctgccca gtgcctcacg accaactttc gctaccttag gaccgttata gttactgcag 3661 cgctagcata acttcgtata aagtatccta tacgaagtta tttgccttaa cccagaaatt 3721 atcactgtta ttctttagaa tggtgcaaag aataacttcg tataatgtat gctatacgaa 3781 gttatgaatt cgatatcaag cttatcgata atcaacctct ggattacaaa atttgtgaaa 3841 gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa 3901 tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat 3961 cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt 4021 gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc 4081 tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc 4141 ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg 4201 ggaaatcatc gtcctttcct tggctgctcg cctatgttgc cacctggatt ctgcgcggga 4261 cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc 4321 tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc 4381 tttgggccgc ctccccgcat cgataccgag cgctgctcga gagatctacg ggtggcatcc 4441 ctgtgacccc tccccagtgc ctctcctggc cctggaagtt gccactccag tgcccaccag 4501 ccttgtccta ataaaattaa gttgcatcat tttgtctgac taggtgtcct tctataatat 4561 tatggggtgg aggggggtgg tatggagcaa ggggcaagtt gggaagacaa cctgtagggc 4621 ctgcggggtc tattgggaac caagctggag tgcagtggca caatcttggc tcactgcaat 4681 ctccgcctcc tgggttcaag cgattctcct gcctcagcct cccgagttgt tgggattcca 4741 ggcatgcatg accaggctca gctaattttt gtttttttgg tagagacggg gtttcaccat 4801 attggccagg ctggtctcca actcctaatc tcaggtgatc tacccacctt ggcctcccaa 4861 attgctggga ttacaggcgt gaaccactgc tcccttccct gtccttctga ttttgtaggt 4921 aaccacgtgc ggaccgagcg gccgcaggaa cccctagtga tggagttggc cactccctct 4981 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt 5041 gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg 5101 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt 5161 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg 5221 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca 5281 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta 5341 gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc 5401 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg 5461 gactcttgtt ccaaactgga acaacactca accctatctc gggctattct tttgatttat 5521 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta 5581 acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct 5641 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct 5701 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct 5761 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga 5821 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca 5881 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata 5941 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga 6001 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc 6061 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg 6121 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc 6181 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat 6241 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact 6301 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat 6361 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga 6421 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc 6481 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga 6541 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag 6601 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc 6661 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt 6721 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct 6781 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg 6841 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg 6901 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca 6961 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga 7021 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa 7081 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga 7141 aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt 7201 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt 7261 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat 7321 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct 7381 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca 7441 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag 7501 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc 7561 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga 7621 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca 7681 tgt //