LOCUS Exported 3004 bp ds-DNA circular SYN 15-JUN-2022 DEFINITION Prime editing in mammalian cells. ACCESSION . VERSION . KEYWORDS pU6-pegRNA-GG-acceptor SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 3004) AUTHORS Anzalone AV, Randolph PB, Davis JR, Sousa AA, Koblan LW, Levy JM, Chen PJ, Wilson C, Newby GA, Raguram A, Liu DR TITLE Search-and-replace genome editing without double-strand breaks or donor DNA. JOURNAL Nature. 2019 Oct 21. pii: 10.1038/s41586-019-1711-4. doi: 10.1038/s41586-019-1711-4. PUBMED 31634902 REFERENCE 2 (bases 1 to 3004) AUTHORS . TITLE Direct Submission JOURNAL Exported Jun 15, 2022 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..3004 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 1..241 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 1..21 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 172..191 /label=LKO.1 5' /note="Human U6 promoter, forward primer" terminator 259..290 /label=tonB terminator /note="bidirectional E. coli tonB-P14 transcription terminator" CDS complement(307..984) /codon_start=1 /product="monomeric derivative of DsRed (Campbell et al., 2002)" /label=mRFP1 /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS TGA" RBS 991..1002 /note="strong bacterial ribosome binding site (Elowitz and Leibler, 2000)" protein_bind 1020..1038 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 1045..1063 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" rep_origin complement(1174..1762) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1254..1273) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(1933..2793) /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 2556..2575 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(2794..2898) /gene="bla" /label=AmpR promoter ORIGIN 1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag 61 ataattagaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga 121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat 181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga 241 cgaaacaccc gagaccgtag tcaaaagcct ccggtcggag gcttttgact tggctgagga 301 gtgccattaa gcaccggtgg agtgacgacc ttcagcacgt tcgtactgtt caacgatggt 361 gtagtcttcg ttgtgggagg tgatgtccag tttgatgtcg gttttgtaag cacccggcag 421 ctgaaccggt tttttagcca tgtaggtggt tttaacttca gcgtcgtagt gaccaccgtc 481 tttcagtttc agacgcattt tgatttcacc tttcagagca ccgtcttccg ggtacatacg 541 ttcggtggaa gcttcccaac ccatggtttt tttctgcata accggaccgt cggacgggaa 601 gttggtacca cgcagtttaa ctttgtagat gaactcaccg tcttgcaggg aggagtcctg 661 ggtaacggta acaacaccac cgtcttcgaa gttcataaca cgttcccatt tgaaaccttc 721 cgggaaggac agtttcaggt agtccgggat gtcagccggg tgtttaacgt aagctttgga 781 accgtactgg aactgcgggg acaggatgtc ccaagcgaac ggcagcggac cacctttggt 841 aactttcagt ttagcggtct gggtaccttc gtacggacga ccttcacctt caccttcgat 901 ttcgaactcg tgaccgttaa cggaaccttc catacgaact ttgaaacgca tgaactcttt 961 gataacgtct tcgctactcg ccatagatcc tttctcctct ttcagatccg tgctcagtat 1021 ctctatcact gatagggatg tcaatctcta tcactgatag ggaggtctcg tttttttaag 1081 cttgggccgc tcgaggtacc tctctacata tgacatgtga gcaaaaggcc agcaaaaggc 1141 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga 1201 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata 1261 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac 1321 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg 1381 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc 1441 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag 1501 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt 1561 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt 1621 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg 1681 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac 1741 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca 1801 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac 1861 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac 1921 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt 1981 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt 2041 accatctggc cccagtgctg caatgatacc gcgagatcca cgctcaccgg ctccagattt 2101 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc 2161 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa 2221 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg 2281 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt 2341 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc 2401 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt 2461 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg 2521 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac 2581 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc 2641 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt 2701 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg 2761 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag 2821 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa 2881 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg ctagctgtac 2941 aaaaaagcag gctttaaagg aaccaattca gtcgactgga tccggtacca aggtcgggca 3001 ggaa //