LOCUS Exported 6666 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION AAV mediated Cre-dependent expression of EGFP (Lox-Stop-Lox) . ACCESSION . VERSION . KEYWORDS pAAV.CAG.LSL.EGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6666) AUTHORS Oh SW, Harris JA, Ng L, Winslow B, Cain N, Mihalas S, Wang Q, Lau C, Kuan L, Henry AM, Mortrud MT, Ouellette B, Nguyen TN, Sorensen SA, Slaughterbeck CR, Wakeman W, Li Y, Feng D, Ho A, Nicholas E, Hirokawa KE, Bohn P, Joines KM, Peng H, Hawrylycz MJ, Phillips JW, Hohmann JG, Wohnoutka P, Gerfen CR, Koch C, Bernard A, Dang C, Jones AR, Zeng H TITLE A mesoscale connectome of the mouse brain. JOURNAL Nature. 2014 Apr 10;508(7495):207-14. doi: 10.1038/nature13186. Epub 2014 Apr 2. PUBMED 24695228 REFERENCE 2 (bases 1 to 6666) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6666 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..130 /label=AAV2 ITR (alternate) /note="Functional equivalent of wild-type AAV2 ITR" enhancer 223..526 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 528..805 /label=chicken beta-actin promoter intron 806..1822 /label=chimeric intron /note="chimera between introns from chicken beta-actin and rabbit beta-globin" primer_bind 1746..1768 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 1830..1849 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" protein_bind 1950..1983 /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)." polyA_signal 2151..2285 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" polyA_signal 2405..2539 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" polyA_signal 2659..2793 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" protein_bind 2822..2855 /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 2890..2899 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2896..3615 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 2896..3612 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(2941..2962) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(3202..3221) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 3549..3570 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(3693..3710) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 3699..3906 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" repeat_region 3929..4069 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 3929..4058 /label=AAV2 ITR rep_origin 4144..4599 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4231..4250) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4441..4462 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4616..4635) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 4735..4757 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(4795..4813) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 4881..4985 /gene="bla" /label=AmpR promoter CDS 4986..5846 /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(5204..5223) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 6017..6605 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6506..6525 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt 61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact 121 aggggttcct gcggccgcac gcgaaacaat tctgcaggaa tctagttatt aatagtaatc 181 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt 241 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta 301 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg 361 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga 421 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt 481 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc 541 cacgttctgc ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt 601 tattttttaa ttattttgtg cagcgatggg ggcggggggg gggggggggc gcgcgccagg 661 cggggcgggg cggggcgagg ggcggggcgg ggcgaggcgg agaggtgcgg cggcagccaa 721 tcagagcggc gcgctccgaa agtttccttt tatggcgagg cggcggcggc ggcggcccta 781 taaaaagcga agcgcgcggc gggcgggagt cgctgcgcgc tgccttcgcc ccgtgccccg 841 ctccgccgcc gcctcgcgcc gcccgccccg gctctgactg accgcgttac tcccacaggt 901 gagcgggcgg gacggccctt ctcctccggg ctgtaattag cgcttggttt aatgacggct 961 tgtttctttt ctgtggctgc gtgaaagcct tgaggggctc cgggagggcc ctttgtgcgg 1021 ggggagcggc tcggggggtg cgtgcgtgtg tgtgtgcgtg gggagcgccg cgtgcggctc 1081 cgcgctgccc ggcggctgtg agcgctgcgg gcgcggcgcg gggctttgtg cgctccgcag 1141 tgtgcgcgag gggagcgcgg ccgggggcgg tgccccgcgg tgcggggggg gctgcgaggg 1201 gaacaaaggc tgcgtgcggg gtgtgtgcgt gggggggtga gcagggggtg tgggcgcgtc 1261 ggtcgggctg caaccccccc tgcacccccc tccccgagtt gctgagcacg gcccggcttc 1321 gggtgcgggg ctccgtacgg ggcgtggcgc ggggctcgcc gtgccgggcg gggggtggcg 1381 gcaggtgggg gtgccgggcg gggcggggcc gcctcgggcc ggggagggct cgggggaggg 1441 gcgcggcggc ccccggagcg ccggcggctg tcgaggcgcg gcgagccgca gccattgcct 1501 tttatggtaa tcgtgcgaga gggcgcaggg acttcctttg tcccaaatct gtgcggagcc 1561 gaaatctggg aggcgccgcc gcaccccctc tagcgggcgc ggggcgaagc ggtgcggcgc 1621 cggcaggaag gaaatgggcg gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct 1681 ccctctccag cctcggggct gtccgcgggg ggacggctgc cttcgggggg gacggggcag 1741 ggcggggttc ggcttctggc gtgtgaccgg cggctctaga gcctctgcta accatgttca 1801 tgccttcttc tttttcctac agctcctggg caacgtgctg gttattgtgc tgtctcatca 1861 ttttggcaaa gaattgattt gataccgcgg gccccgcgtg gacggtaccc ttcaggtcga 1921 catttatccc taaacaggat aacactagca taacttcgta taggatactt tatacgaagt 1981 tattcgcgat gaataaatga aagcttgcag atctgcgact ctagaggatc tgcgactcta 2041 gaggatcata atcagccata ccacattttg tagaggtttt acttgcttta aaaaacctcc 2101 cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta 2161 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat 2221 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct 2281 ggatctgcga ctctagagga tcataatcag ccataccaca tttgtagagg ttttacttgc 2341 tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt 2401 tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt 2461 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt 2521 atcttatcat gtctggatct gcgactctag aggatcataa tcagccatac cacatttgta 2581 gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg 2641 aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat 2701 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc 2761 aaactcatca atgtatctta tcatgtctgg atccccatca agctgatccg gaacccttaa 2821 tataacttcg tataggatac tttatacgaa gttattagct agttacagta ccggcttagt 2881 taggaattcg ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc 2941 ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag 3001 ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc 3061 gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac 3121 cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag 3181 gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc 3241 gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc 3301 aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc 3361 gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc 3421 agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg 3481 ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag 3541 cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac 3601 gagctgtaca agtaagaatt ctaaatttcg aaagtcgacg tcggttggcg cgccaagggt 3661 ttaaaccccc tggccggcct tgcgaagatc tgcctcgact gtgccttcta gttgccagcc 3721 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt 3781 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct 3841 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc 3901 tggggacacg tgcggaccga gcggccgcag gaacccctag tgatggagtt ggccactccc 3961 tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc 4021 tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg 4081 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat 4141 agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga 4201 ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg 4261 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat 4321 ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg 4381 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata 4441 gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt 4501 tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat 4561 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa 4621 tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc 4681 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga 4741 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg 4801 tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg 4861 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa 4921 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga 4981 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc 5041 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg 5101 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc 5161 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat 5221 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg 5281 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag 5341 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa 5401 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc 5461 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca 5521 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc 5581 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc 5641 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg 5701 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta 5761 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag 5821 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga 5881 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc 5941 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa 6001 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa 6061 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc 6121 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt 6181 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc 6241 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac 6301 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca 6361 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg 6421 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag 6481 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt 6541 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat 6601 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc 6661 acatgt //