LOCUS       sequence-352079-        6556 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of base editor .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6556)
  AUTHORS   Mok BY, Kotrys AV, Raguram A, Huang TP, Mootha VK, Liu DR
  TITLE     CRISPR-free base editors with enhanced activity and expanded 
            targeting scope in mitochondrial and nuclear DNA.
  JOURNAL   Nat Biotechnol. 2022 Sep;40(9):1378-1387. doi: 
            10.1038/s41587-022-01256-8. Epub 2022 Apr 4.
  PUBMED    35379961
REFERENCE   2  (bases 1 to 6556)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41587-022-01256-8"; journalName: "Nat Biotechnol"; date: 
            "2022-09"; volume: "40"; issue: "9"; pages: "1378-1387"
COMMENT     Sequence Label: pBM19-sd4U
FEATURES             Location/Qualifiers
     source          1..6556
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(58..873)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(58..873)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     781..800
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     865..884
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(1333..1350)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1467..2012)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(1547..1566)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2088..2107
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     terminator      2300..2345
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2360..3238)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        3265..3549
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     3472..3491
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     RBS             3678..3686
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     primer_bind     complement(5624..5641)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      5794..5880
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5972..5999
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 acttattcag gcgtagcaac caggcgttta agggcaccaa taactgcctt aaaaaaatta
       61 gaaaaactca tcgagcatca aatgaaactg caatttattc atatcaggat tatcaatacc
      121 atatttttga aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc agttccatag
      181 gatggcaaga tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa tacaacctat
      241 taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag tgacgactga
      301 atccggtgag aatggcaaaa gtttatgcat ttctttccag acttgttcaa caggccagcc
      361 attacgctcg tcatcaaaat cactcgcatc aaccaaaccg ttattcattc gtgattgcgc
      421 ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg
      481 caaccggcgc aggaacactg ccagcgcatc aacaatattt tcacctgaat caggatattc
      541 ttctaatacc tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc
      601 aggagtacgg ataaaatgct tgatggtcgg aagaggcata aattccgtca gccagtttag
      661 tctgaccatc tcatctgtaa catcattggc aacgctacct ttgccatgtt tcagaaacaa
      721 ctctggcgca tcgggcttcc catacaatcg atagattgtc gcacctgatt gcccgacatt
      781 atcgcgagcc catttatacc catataaatc agcatccatg ttggaattta atcgcggcct
      841 agagcaagac gtttcccgtt gaatatggct cataacaccc cttgtattac tgtttatgta
      901 agcagacagt tttattgttc atgaagagaa acctacggat cacgtctcat tttcgccaaa
      961 agttggccca gggcttcccg gtatcaacag ggacaccagg atttatttat tctgcgaagt
     1021 gatcttccgt cacaggtatt tattcggcgc aaagtgcgtc gggtgatgct gccaacttac
     1081 tgatttagtg tatgatggtg tttttgaggt gctccagtgg cttctgtttc tatcagctgt
     1141 ccctcctgtt cagctactga cggggtggtg cgtaacggca aaagcaccgc cggacatcag
     1201 cgctagcgga gtgtatactg gcttactatg ttggcactga tgagggtgtc agtgaagtgc
     1261 ttcatgtggc aggagaaaaa aggctgcacc ggtgcgtcag cagaatatgt gatacaggat
     1321 atattccgct tcctcgctca ctgactcgct acgctcggtc gttcgactgc ggcgagcgga
     1381 aatggcttac gaacggggcg gagatttcct ggaagatgcc aggaagatac ttaacaggga
     1441 agtgagaggg ccgcggcaaa gccgtttttc cataggctcc gcccccctga caagcatcac
     1501 gaaatctgac gctcaaatca gtggtggcga aacccgacag gactataaag ataccaggcg
     1561 tttccccctg gcggctccct cgtgcgctct cctgttcctg cctttcggtt taccggtgtc
     1621 attccgctgt tatggccgcg tttgtctcat tccacgcctg acactcagtt ccgggtaggc
     1681 agttcgctcc aagctggact gtatgcacga accccccgtt cagtccgacc gctgcgcctt
     1741 atccggtaac tatcgtcttg agtccaaccc ggaaagacat gcaaaagcac cactggcagc
     1801 agccactggt aattgattta gaggagttag tcttgaagtc atgcgccggt taaggctaaa
     1861 ctgaaaggac aagttttggt gactgcgctc ctccaagcca gttacctcgg ttcaaagagt
     1921 tggtagctca gagaacattc gaaaaaccgc cctgcaaggc ggttttttcg ttttcagagc
     1981 aagagattac gcgcagacca aaacgatctc aagaagatca tcttattaat cagataaaat
     2041 atttctaggc tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg
     2101 atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg
     2161 tagaggatct gctcatgttt gacagcttat catcgatgca taatgtgcct gtcaaatgga
     2221 cgaagcaggg attctgcaaa ccctatgcta ctccgtcaag ccgtcaattg tctgattcgt
     2281 taccaaactt agggactctc aaataaaacg aaaggctcag tcgaaagact gggcctttcg
     2341 ttttagactt agggaccctt tatgacaact tgacggctac atcattcact ttttcttcac
     2401 aaccggcacg gaactcgctc gggctggccc cggtgcattt tttaaatacc cgcgagaaat
     2461 agagttgatc gtcaaaacca acattgcgac cgacggtggc gataggcatc cgggtggtgc
     2521 tcaaaagcag cttcgcctgg ctgatacgtt ggtcctcgcg ccagcttaag acgctaatcc
     2581 ctaactgctg gcggaaaaga tgtgacagac gcgacggcga caagcaaaca tgctgtgcga
     2641 cgctggcgat atcaaaattg ctgtctgcca ggtgatcgct gatgtactga caagcctcgc
     2701 gtacccgatt atccatcggt ggatggagcg actcgttaat cgcttccatg cgccgcagta
     2761 acaattgctc aagcagattt atcgccagca gctccgaata gcgcccttcc ccttgcccgg
     2821 cgttaatgat ttgcccaaac aggtcgctga aatgcggctg gtgcgcttca tccgggcgaa
     2881 agaaccccgt attggcaaat attgacggcc agttaagcca ttcatgccag taggcgcgcg
     2941 gacgaaagta aacccactgg tgataccatt cgcgagcctc cggatgacga ccgtagtgat
     3001 gaatctctcc tggcgggaac agcaaaatat cacccggtcg gcaaacaaat tctcgtccct
     3061 gatttttcac caccccctga ccgcgaatgg tgagattgag aatataacct ttcattccca
     3121 gcggtcggtc gataaaaaaa tcgagataac cgttggcctc aatcggcgtt aaacccgcca
     3181 ccagatgggc attaaacgag tatcccggca gcaggggatc attttgcgct tcagccatac
     3241 ttttcatact cccaccattc agagaagaaa ccaattgtcc atattgcatc agacattgcc
     3301 gtcactgcgt cttttactgg ctcttctcgc taacccaacc ggtaaccccg cttattaaaa
     3361 gcattctgta acaaagcggg accaaagcca tgacaaaaac gcgtaacaaa agtgtctata
     3421 atcacggcag aaaagtccac attgattatt tgcacggcgt cacactttgc tatgccatag
     3481 catttttatc cataagatta gcggatccta cctgacgctt tttatcgcaa ctctctactg
     3541 tttctccata cccgtttttt tggacgcgta caactcaagt ctgacaaaaa aaaggaaaaa
     3601 ttgactaatc tgagcgacat cattgagaag gagactggga aacagctggt cattcaggag
     3661 tccatcctga tgctgcctaa ggaggtgaag gaagtgatcg gcaacaagcc agagtctgac
     3721 atcctggtgc acaccgccta cgacgagtcc acagatgaga atgtgatgct gctgacctct
     3781 gacgcccccg agtataagcc ttgggccctg gtcatccagg attctaacgg cgagaataag
     3841 atcaagatgc tgtctggtgg ttctaacatc gccgatctgc ggaccctggg ctactcccag
     3901 cagcagcagg agaagatcaa gccaaaggtg cgctctaccg tggctcagca ccacgaggcc
     3961 ctggtgggac acggcttcac acacgctcac atcgtggccc tgtcccagca ccctgccgct
     4021 ctgggcaccg tggccgtgaa gtaccaggac atgatcgccg ctctgccaga ggctacacac
     4081 gaggccatcg tgggagtggg caagcagtgg tctggagctc gggctctgga ggctctgctg
     4141 accgtggctg gagagctaag ggaaccacct ctgcagctgg ataccggcca gctgctgaag
     4201 atcgccaaga ggggaggagt gacagctgtg gaggctgtgc acgcttggag aaacgctctg
     4261 accggagctc ctctgaacct gaccccagaa caggttgtgg ctatcgcctc tcacgacggc
     4321 ggcaaacagg ccctggaaac cgtgcaggct ctgctgccag tgctgtgtca ggcccacggc
     4381 ctgacccctc agcaggtggt ggccatagcc agcaacaacg gcggcaaaca ggccctggaa
     4441 accgtgcagc ggctgctgcc cgtgctgtgc caagcccacg gccttacacc acagcaggtg
     4501 gtggctatcg ccagcaacgg cggcggcaag caggctctgg aaaccgttca gcggctgctg
     4561 cctgtgctgt gtcaagccca cggcctgacc cccgagcagg tggtggccat cgcctcccac
     4621 gacgggggca agcaggccct ggagacagtg caggcccttc tgcctgtgct gtgccaagcc
     4681 cacggcctga cacccgagca ggtggtggcc attgcttcta acaacggcgg aaagcaggcc
     4741 ctggagacag tgcaggccct gctgcctgtg ctgtgccagg ctcatggcct gaccccagag
     4801 caggtggtgg ccatcgcctc caatggaggc ggcaagcagg ccctggaaac cgtgcaggcc
     4861 ctgctgcctg tgctgtgcca ggcccacgga ctgacacccg agcaagtggt cgccatcgct
     4921 agcaacggcg gcggaaagca ggctctggaa accgttcaaa gactgctgcc cgtgctgtgc
     4981 caggctcacg gcctgacccc tgagcaggtc gtggccatcg ccagcaatgg cggaggcaag
     5041 caggctctgg agacagtgca ggccctgctg cccgtcctgt gccaggccca tggcctcacc
     5101 cctgagcagg tggtggccat tgccagcaac aacggcggaa agcaggccct ggaaaccgtg
     5161 cagagactgc tgcctgtgct gtgccaggca cacggcctga cccctgagca ggtggtggcc
     5221 atcgccagcc acgatggcgg aaagcaagct ctggaaaccg tgcaggccct gctgcccgtg
     5281 ctgtgtcagg cccacggcct gacacctcaa caagtagttg ccatcgccag caacggcggc
     5341 ggtagacctg ccctcgagtc tatcgtggct cagctgagcc gcccaaaccc cgccctggcc
     5401 gctctgacca acgatcgcct ggtggccctg gcttgtctgg gaggccggcc tgctctgaac
     5461 gccgtgaaga agggactggg cggatccgcc attccagtga agcgcggcgc taccggtgaa
     5521 accaaagtgt ttatcggtaa cagcaacagc ccgaagagcc cgaccaaagg cggttgctaa
     5581 tcgacaagct tggctgtttt ggcggatgag agaagatttt cagcctgata cagattaaat
     5641 cagaacgcag aagcggtctg ataaaacaga atttgcctgg cggcagtagc gcggtggtcc
     5701 cacctgaccc catgccgaac tcagaagtga aacgccgtag cgccgatggt agtgtggggt
     5761 ctccccatgc gagagtaggg aactgccagg catcaaataa aacgaaaggc tcagtcgaaa
     5821 gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag taggacaaat
     5881 ccgccgggag cggatttgaa cgttgcgaag caacggcccg gagggtggcg ggcaggacgc
     5941 ccgccataaa ctgccaggca tcaaattaag cagaaggcca tcctgacgga tggccttttt
     6001 gcgtttctac aaactctctg tcagaccaag tttactcata tatactttag attgatttaa
     6061 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     6121 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     6181 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     6241 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     6301 gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     6361 agcaccgcct acatacctcg ctctgctaat cctgttacca gtcaggcatt tgagaagcac
     6421 acggtcacac tgcttccggt agtcaataaa ccggtaaacc agcaatagac ataagcggct
     6481 atttaacgac cctgccctga accgacgacc gggtcgaatt tgctttcgaa tttctgccat
     6541 tcatccgctt attatc
//
