LOCUS Exported 7378 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION hu6 HGPS sgRNA expression and ABE7.10max VRQR C terminal AAV vector. ACCESSION . VERSION . KEYWORDS hu6 HGPS sgRNA expression and ABE7.10max VRQR C terminal AAV v... SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7378) AUTHORS Koblan LW, Erdos MR, Wilson C, Cabral WA, Levy JM, Xiong ZM, Tavarez UL, Davison LM, Gete YG, Mao X, Newby GA, Doherty SP, Narisu N, Sheng Q, Krilow C, Lin CY, Gordon LB, Cao K, Collins FS, Brown JD, Liu DR TITLE In vivo base editing rescues Hutchinson-Gilford progeria syndrome in mice. JOURNAL Nature. 2021 Jan;589(7843):608-614. doi: 10.1038/s41586-020-03086-7. Epub 2021 Jan 6. PUBMED 33408413 REFERENCE 2 (bases 1 to 7378) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7378 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 1..286 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer; contains an 18-bp deletion relative to the standard CMV enhancer" promoter 288..565 /label=chicken beta-actin promoter intron 566..793 /label=hybrid intron /note="hybrid between chicken beta-actin (CBA) and minute virus of mice (MMV) introns (Gray et al., 2011)" CDS 847..867 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 3400..3420 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" primer_bind complement(3485..3505) /label=WPRE-R /note="WPRE, reverse primer" polyA_signal 3681..3905 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" misc_RNA complement(3919..3994) /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" promoter complement(4023..4263) /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind complement(4073..4092) /label=LKO.1 5' /note="Human U6 promoter, forward primer" primer_bind complement(4243..4263) /label=hU6-F /note="Human U6 promoter, forward primer" repeat_region 4287..4416 /label=AAV2 ITR primer_bind complement(4435..4452) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(4435..4451) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4444..4466) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 4593..5048 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4680..4699) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4890..4911 /label=F1ori-F /note="F1 origin, forward primer" promoter 5074..5178 /gene="bla" /label=AmpR promoter CDS 5179..6039 /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(5397..5416) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 6210..6798 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6699..6718 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 6952..6969 /label=L4440 /note="L4440 vector, forward primer" protein_bind 7086..7107 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 7122..7152 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 7160..7176 /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 7165..7187 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 7184..7200 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 7184..7200 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" repeat_region 7227..7356 /label=AAV2 ITR ORIGIN 1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt 61 gacgtcaata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg 121 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga 181 cgtcaatgac ggtaaatggc ccgcctggca ttgtgcccag tacatgacct tatgggactt 241 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc 301 cacgttctgc ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt 361 tattttttaa ttattttgtg cagcgatggg ggcggggggg gggggggggc gcgcgccagg 421 cggggcgggg cggggcgagg ggcggggcgg ggcgaggcgg agaggtgcgg cggcagccaa 481 tcagagcggc gcgctccgaa agtttccttt tatggcgagg cggcggcggc ggcggcccta 541 taaaaagcga agcgcgcggc gggcgggagt cgctgcgcgc tgccttcgcc ccgtgccccg 601 ctccgccgcc gcctcgcgcc gcccgccccg gctctgactg accgcgttac tcccacaggt 661 gagcgggcgg gacggccctt ctcctccggg ctgtaattag ctgagcaaga ggtaagggtt 721 taagggatgg ttggttggtg gggtattaat gtttaattac ctggagcacc tgcctgaaat 781 cacttttttt caggttggac cggtgccacc atgaaacgga cagccgacgg aagcgagttc 841 gagtcaccaa agaagaagcg gaaagtcatc aagattgcta cacggaaata cctgggaaag 901 cagaacgtgt acgacatcgg cgtggagcgg gatcacaact tcgccctgaa gaatggcttt 961 atcgccagca attgcttcga ctccgtggaa atctccggcg tggaagatcg gttcaacgcc 1021 tccctgggca cataccacga tctgctgaaa attatcaagg acaaggactt cctggacaat 1081 gaggaaaacg aggacattct ggaagatatc gtgctgaccc tgacactgtt tgaggacaga 1141 gagatgatcg aggaacggct gaaaacctat gcccacctgt tcgacgacaa agtgatgaag 1201 cagctgaagc ggcggagata caccggctgg ggcaggctga gccggaagct gatcaacggc 1261 atccgggaca agcagtccgg caagacaatc ctggatttcc tgaagtccga cggcttcgcc 1321 aacagaaact tcatgcagct gatccacgac gacagcctga cctttaaaga ggacatccag 1381 aaagcccagg tgtccggcca gggcgatagc ctgcacgagc acattgccaa tctggccggc 1441 agccccgcca ttaagaaggg catcctgcag acagtgaagg tggtggacga gctcgtgaaa 1501 gtgatgggcc ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga gaaccagacc 1561 acccagaagg gacagaagaa cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa 1621 gagctgggca gccagatcct gaaagaacac cccgtggaaa acacccagct gcagaacgag 1681 aagctgtacc tgtactacct gcagaatggg cgggatatgt acgtggacca ggaactggac 1741 atcaaccggc tgtccgacta cgatgtggac catatcgtgc ctcagagctt tctgaaggac 1801 gactccatcg ataacaaggt gctgaccaga agcgacaaga accggggcaa gagcgacaac 1861 gtgccctccg aagaggtcgt gaagaagatg aagaactact ggcggcagct gctgaacgcc 1921 aagctgatta cccagagaaa gttcgacaat ctgaccaagg ccgagagagg cggcctgagc 1981 gaactggata aggccggctt catcaagaga cagctggtgg aaacccggca gatcacaaag 2041 cacgtggcac agatcctgga ctcccggatg aacactaagt acgacgagaa tgacaagctg 2101 atccgggaag tgaaagtgat caccctgaag tccaagctgg tgtccgattt ccggaaggat 2161 ttccagtttt acaaagtgcg cgagatcaac aactaccacc acgcccacga cgcctacctg 2221 aacgccgtcg tgggaaccgc cctgatcaaa aagtacccta agctggaaag cgagttcgtg 2281 tacggcgact acaaggtgta cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc 2341 ggcaaggcta ccgccaagta cttcttctac agcaacatca tgaacttttt caagaccgag 2401 attaccctgg ccaacggcga gatccggaag cggcctctga tcgagacaaa cggcgaaacc 2461 ggggagatcg tgtgggataa gggccgggat tttgccaccg tgcggaaagt gctgagcatg 2521 ccccaagtga atatcgtgaa aaagaccgag gtgcagacag gcggcttcag caaagagtct 2581 atcctgccca agaggaacag cgataagctg atcgccagaa agaaggactg ggaccctaag 2641 aagtacggcg gcttcgtgag ccccaccgtg gcctattctg tgctggtggt ggccaaagtg 2701 gaaaagggca agtccaagaa actgaagagt gtgaaagagc tgctggggat caccatcatg 2761 gaaagaagca gcttcgagaa gaatcccatc gactttctgg aagccaaggg ctacaaagaa 2821 gtgaaaaagg acctgatcat caagctgcct aagtactccc tgttcgagct ggaaaacggc 2881 cggaagagaa tgctggcctc agccagagaa ctgcagaagg gaaacgaact ggccctgccc 2941 tccaaatatg tgaacttcct gtacctggcc agccactatg agaagctgaa gggctccccc 3001 gaggataatg agcagaaaca gctgtttgtg gaacagcaca agcactacct ggacgagatc 3061 atcgagcaga tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa 3121 gtgctgtccg cctacaacaa gcaccgggat aagcccatca gagagcaggc cgagaatatc 3181 atccacctgt ttaccctgac caatctggga gcccctgccg ccttcaagta ctttgacacc 3241 accatcgacc ggaagcagta cagaagcacc aaagaggtgc tggacgccac cctgatccac 3301 cagagcatca ccggcctgta cgagacacgg atcgacctgt ctcagctggg aggtgactct 3361 ggcggctcaa aaagaaccgc cgacggcagc gaattcgagc ccaagaagaa gaggaaagtc 3421 taagatctga taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta 3481 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta 3541 ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggtta gttcttgcca 3601 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca 3661 ctgacaattc cgtggtgcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc 3721 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga 3781 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca 3841 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc 3901 tatggctcga gaaaaaaagc accgactcgg tgccactttt tcaagttgat aacggactag 3961 ccttatttta acttgctatt tctagctcta aaacggagcc caggtgggtg gaccggtgtt 4021 tcgtcctttc cacaagatat ataaagccaa gaaatcgaaa tactttcaag ttacggtaag 4081 catatgatag tccattttaa aacataattt taaaactgca aactacccaa gaaattatta 4141 ctttctacgt cacgtatttt gtactaatat ctttgtgttt acagtcaaat taattctaat 4201 tatctctcta acagccttgt atcgtatatg caaatatgaa ggaatcatgg gaaataggcc 4261 ctcttcctgc ccgaccttgc ggccgcagga acccctagtg atggagttgg ccactccctc 4321 tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt 4381 tgcccgggcg gcctcagtga gcgagcgagc gcgcagcctt aattaaccta attcactggc 4441 cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc 4501 agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc 4561 ccaacagttg cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc 4621 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc 4681 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct 4741 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa 4801 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc 4861 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact 4921 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg 4981 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct 5041 tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc 5101 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa 5161 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt 5221 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct 5281 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc 5341 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta 5401 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac 5461 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc 5521 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac 5581 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg 5641 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac 5701 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc 5761 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt 5821 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga 5881 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc 5941 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag 6001 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca 6061 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc 6121 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca 6181 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc 6241 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta 6301 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt 6361 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc 6421 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg 6481 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg 6541 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag 6601 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc 6661 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat 6721 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg 6781 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc 6841 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt 6901 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca 6961 gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg 7021 attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac 7081 gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg 7141 gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac 7201 catgattacg ccagatttaa ttaaggctgc gcgctcgctc gctcactgag gccgcccggg 7261 caaagcccgg gcgtcgggcg acctttggtc gcccggcctc agtgagcgag cgagcgcgca 7321 gagagggagt ggccaactcc atcactaggg gttcctgcgg cctctagatc agggtacc //