LOCUS Exported 3041 bp ds-DNA circular SYN 15-AUG-2023 DEFINITION Prime editing in mammalian cells. ACCESSION . VERSION . KEYWORDS pU6-tevopreq1-GG-acceptor SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 3041) AUTHORS Nelson JW, Randolph PB, Shen SP, Everette KA, Chen PJ, Anzalone AV, An M, Newby GA, Chen JC, Hsu A, Liu DR TITLE Engineered pegRNAs improve prime editing efficiency. JOURNAL Nat Biotechnol. 2021 Oct 4. pii: 10.1038/s41587-021-01039-7. doi: 10.1038/s41587-021-01039-7. PUBMED 34608327 REFERENCE 2 (bases 1 to 3041) AUTHORS . TITLE Direct Submission JOURNAL Exported Aug 15, 2023 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..3041 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 82..322 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 82..102 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 253..272 /label=LKO.1 5' /note="Human U6 promoter, forward primer" terminator 340..371 /label=tonB terminator /note="bidirectional E. coli tonB-P14 transcription terminator" CDS complement(388..1065) /codon_start=1 /product="monomeric derivative of DsRed (Campbell et al., 2002)" /label=mRFP1 /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS TGA" RBS 1072..1083 /note="strong bacterial ribosome binding site (Elowitz and Leibler, 2000)" protein_bind 1101..1119 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 1126..1144 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" rep_origin complement(1292..1880) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1372..1391) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(2051..2911) /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 2674..2693 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(2912..3016) /gene="bla" /label=AmpR promoter ORIGIN 1 gacgtcgcta gctgtacaaa aaagcaggct ttaaaggaac caattcagtc gactggatcc 61 ggtaccaagg tcgggcagga agagggccta tttcccatga ttccttcata tttgcatata 121 cgatacaagg ctgttagaga gataattaga attaatttga ctgtaaacac aaagatatta 181 gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt tttaaaatta 241 tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga tttcttggct 301 ttatatatct tgtggaaagg acgaaacacc cgagaccgta gtcaaaagcc tccggtcgga 361 ggcttttgac ttggctgagg agtgccatta agcaccggtg gagtgacgac cttcagcacg 421 ttcgtactgt tcaacgatgg tgtagtcttc gttgtgggag gtgatgtcca gtttgatgtc 481 ggttttgtaa gcacccggca gctgaaccgg ttttttagcc atgtaggtgg ttttaacttc 541 agcgtcgtag tgaccaccgt ctttcagttt cagacgcatt ttgatttcac ctttcagagc 601 accgtcttcc gggtacatac gttcggtgga agcttcccaa cccatggttt ttttctgcat 661 aaccggaccg tcggacggga agttggtacc acgcagttta actttgtaga tgaactcacc 721 gtcttgcagg gaggagtcct gggtaacggt aacaacacca ccgtcttcga agttcataac 781 acgttcccat ttgaaacctt ccgggaagga cagtttcagg tagtccggga tgtcagccgg 841 gtgtttaacg taagctttgg aaccgtactg gaactgcggg gacaggatgt cccaagcgaa 901 cggcagcgga ccacctttgg taactttcag tttagcggtc tgggtacctt cgtacggacg 961 accttcacct tcaccttcga tttcgaactc gtgaccgtta acggaacctt ccatacgaac 1021 tttgaaacgc atgaactctt tgataacgtc ttcgctactc gccatagatc ctttctcctc 1081 tttcagatcc gtgctcagta tctctatcac tgatagggat gtcaatctct atcactgata 1141 gggaggtctc gcgcggttct atctagttac gcgttaaacc aactagaatt tttttaagct 1201 tgggccgctc gaggtacctc tctacatatg acatgtgagc aaaaggccag caaaaggcca 1261 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc 1321 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc 1381 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg 1441 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta 1501 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg 1561 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac 1621 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag 1681 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat 1741 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat 1801 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc 1861 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt 1921 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct 1981 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt 2041 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc 2101 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac 2161 catctggccc cagtgctgca atgataccgc gagatccacg ctcaccggct ccagatttat 2221 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg 2281 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata 2341 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta 2401 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt 2461 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag 2521 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa 2581 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc 2641 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt 2701 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc 2761 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta 2821 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa 2881 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca 2941 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac 3001 aaataggggt tccgcgcaca tttccccgaa aagtgccacc t //