LOCUS Exported 5911 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION AAV expression of Ruby2 spaghetti monster FLAG construct. ACCESSION . VERSION . KEYWORDS pAAV.CAG.Ruby2sm-Flag.WPRE.SV40 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5911) AUTHORS Viswanathan S, Williams ME, Bloss EB, Stasevich TJ, Speer CM, Nern A, Pfeiffer BD, Hooks BM, Li WP, English BP, Tian T, Henry GL, Macklin JJ, Patel R, Gerfen CR, Zhuang X, Wang Y, Rubin GM, Looger LL TITLE High-performance probes for light and electron microscopy. JOURNAL Nat Methods. 2015 Jun;12(6):568-76. doi: 10.1038/nmeth.3365. Epub 2015 Apr 27. PUBMED 25915120 REFERENCE 2 (bases 1 to 5911) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5911 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..130 /label=AAV2 ITR promoter 441..718 /label=chicken beta-actin promoter primer_bind 997..1019 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 1081..1100 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" regulatory 1144..1153 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1153..1176 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 1180..1203 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 1207..1230 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 1735..1758 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 1765..1788 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 1801..1824 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 1831..1854 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 2047..2070 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 2074..2097 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 2101..2124 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" misc_feature 2152..2740 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2205..2225) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(2623..2634) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 2783..2904 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2794..2813) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2848..2867 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 3066..3195 /label=AAV2 ITR rep_origin 3267..3722 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3354..3373) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3564..3585 /label=F1ori-F /note="F1 origin, forward primer" promoter 3748..3852 /gene="bla" /label=AmpR promoter CDS 3853..4713 /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(4071..4090) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 4884..5472 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5373..5392 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5626..5643 /label=L4440 /note="L4440 vector, forward primer" protein_bind 5760..5781 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 5796..5826 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5834..5850 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." primer_bind 5839..5861 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 5858..5874 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 5858..5874 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" ORIGIN 1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt 61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact 121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct 181 aggaagagta ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac 241 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca 301 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg 361 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt 421 agtcatcgct attaccatgg tcgaggtgag ccccacgttc tgcttcactc tccccatctc 481 ccccccctcc ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat 541 gggggcgggg gggggggggg ggcgcgcgcc aggcggggcg gggcggggcg aggggcgggg 601 cggggcgagg cggagaggtg cggcggcagc caatcagagc ggcgcgctcc gaaagtttcc 661 ttttatggcg aggcggcggc ggcggcggcc ctataaaaag cgaagcgcgc ggcgggcggg 721 agtcgctgcg cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc gccgcccgcc 781 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc 841 gggctgtaat tagcgcttgg tttaatgacg gcttgtttct tttctgtggc tgcgtgaaag 901 ccttgagggg ctccgggagg gccctttgtg cggggggagc ggctcggggc tgtccgcggg 961 gggacggctg ccttcggggg ggacggggca gggcggggtt cggcttctgg cgtgtgaccg 1021 gcggctctag agcctctgct aaccatgttc atgccttctt ctttttccta cagctcctgg 1081 gcaacgtgct ggttattgtg ctgtctcatc attttggcaa agaattggat ccgcggccgc 1141 gccgccacca tggactacaa ggacgacgac gacaaaggtg actacaaaga tgatgacgat 1201 aaaggcgact ataaggacga tgacgacaag ggcggaaact cactgatcaa ggaaaacatg 1261 cggatgaagg tggtgatgga gggctccgtg aatggtcacc agttcaagtg caccggagag 1321 ggagagggaa acccgtacat gggaactcag accatgcgca ttaaggtcat cgaaggaggt 1381 ccgctgccgt tcgctttcga tatcctggcc acttcgttcg gaggagggtc gcgcacgttc 1441 atcaagtacc cgaagggaat cccggacttc tttaagcagt cattcccgga aggattcact 1501 tgggaacggg tgacccggta tgaagatgga ggtgtggtga ctgtcatgca agatacttcg 1561 ctggaggatg ggtgcctcgt gtaccacgtc caagtccgcg gagtgaattt cccgtccaac 1621 ggaccagtga tgcagaaaaa gacgaagggt tgggaaccta atactgaaat gatgtacccc 1681 gcagacggag ggctgagggg ctacacccac atggcgctga aggtcgacgg aggagattac 1741 aaggatgacg acgataagca acaagattac aaagacgatg atgacaaggg ccagcagggc 1801 gactacaagg acgacgacga caagcagcag gactacaaag atgacgatga taaaggagga 1861 ggacatctgt cctgttcgtt cgtgaccacc tacagatcaa agaaaaccgt gggaaacatc 1921 aagatgccgg gcattcatgc cgtcgaccac cgcctggagc ggctcgaaga atcagacaat 1981 gagatgttcg tcgtgcaaag agaacatgcc gtggccaagt tcgcgggact gggaggcggt 2041 ggaggcgatt acaaagacga tgatgacaag ggtgactata aagacgacga tgacaaaggg 2101 gattacaagg atgatgatga taagtagacc ggtgaattca agcttatcga taatcaacct 2161 ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg 2221 ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc 2281 attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt 2341 gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc 2401 attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg 2461 gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact 2521 gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt 2581 gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg 2641 gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc 2701 cctcagacga gtcggatctc cctttgggcc gcctccccgc atcgataccg tcgacccggg 2761 cggccgcttc gagcagacat gataagatac attgatgagt ttggacaaac cacaactaga 2821 atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc 2881 attataagct gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt 2941 cagggggaga tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtaaaatc 3001 gataaggatc ttcctagagc atggctacgt agataagtag catggcgggt taatcattaa 3061 ctacaaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac 3121 tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag 3181 cgagcgagcg cgcagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca 3241 gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg 3301 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt 3361 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg 3421 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga 3481 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac 3541 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc 3601 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa 3661 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat 3721 ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata 3781 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga 3841 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca 3901 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat 3961 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag 4021 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc 4081 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct 4141 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca 4201 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt 4261 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat 4321 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt 4381 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta 4441 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga 4501 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt 4561 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc 4621 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct 4681 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata 4741 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt 4801 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc 4861 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg 4921 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact 4981 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg 5041 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg 5101 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac 5161 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca 5221 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga 5281 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc 5341 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct 5401 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg 5461 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct 5521 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc 5581 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc 5641 gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat 5701 taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt 5761 aatgtgagtt agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt 5821 atgttgtgtg gaattgtgag cggataacaa tttcacacag gaaacagcta tgaccatgat 5881 tacgccagat ttaattaagg ccttaattag g //