LOCUS       sequence-351813-        7238 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression  of base editor in mammalian cells .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7238)
  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 7238)
  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: ND4.2-Left TALE-G1397-N-wildtype
FEATURES             Location/Qualifiers
     source          1..7238
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(80..99)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        271..650
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        651..854
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     805..825
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     899..918
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        899..917
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             940..1017
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MASVLTPLLLRGLTGSARRLPVPRAK"
     CDS             1030..1095
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3058..3309
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     rep_origin      3506..3934
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3593..3612)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3803..3824
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3943..3963)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3948..4277
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4128..4263
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4190..4209
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4325..4372
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4391..4789
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    4947..5068
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4984..5003)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5038..5057
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5117..5133)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5117..5133)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5130..5152)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5141..5157
                     /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)."
     promoter        complement(join(5165..5171,5172..5189,5190..5195))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5210..5231
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5348..5365)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5519..6107)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5599..5618)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6278..7069,7070..7138))
                     /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     6901..6920
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7139..7238,1..5))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc
       61 ccctatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc
      121 tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa
      181 ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc
      241 ttcgcgatgt acgggccaga tatacgcgtt gacattgatt attgactagt tattaatagt
      301 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
      361 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
      421 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
      481 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
      541 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
      601 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt
      661 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
      721 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
      781 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
      841 atataagcag agctctctgg ctaactagag aacccactgc ttactggctt atcgaaatta
      901 atacgactca ctatagggag acccaagctg gctagcacca tggcctccgt tctgaccccg
      961 ctgctgctgc gtggcctgac cggctccgct cgtcgtctgc cagttccgcg tgcgaaaatc
     1021 cattccctgg actacaaaga ccatgacggt gattataaag atcatgacat cgattacaag
     1081 gatgacgatg acaagatgga catcgcggat ctgcgtaccc tgggttacag ccagcagcag
     1141 caggagaaga tcaagccgaa ggtgcgcagc accgtggctc agcaccacga agccctggtg
     1201 ggccacggtt tcacccacgc tcacatcgtg gccctgagcc agcacccagc cgcgctgggc
     1261 accgtggccg tgaaatatca ggacatgatt gctgccctgc cagaggccac ccatgaagct
     1321 attgtgggcg tgggcaagca gtggagcggt gctcgtgcgc tggaggcgct gctgaccgtg
     1381 gctggtgaac tgcgtggtcc gccgctgcag ctggataccg gtcagctgct gaaaattgcc
     1441 aaacgtggcg gtgtgaccgc ggtggaagcc gtgcatgctt ggcgtaatgc tctgaccggt
     1501 gcgccgctga acttaacgcc ggaacaagtt gttgcaattg ccagcaacaa cggcggcaag
     1561 caagctctgg aaaccgtgca ggccctgctt ccagtcctgt gtcaggccca cggcctgacc
     1621 cctcagcagg tggtcgccat cgcctctaac atcggcggaa agcaggctct ggagaccgtg
     1681 cagcggctgc tgcccgtgct gtgccaggcc cacggcctga cacctcagca ggtggtggcc
     1741 attgctagca acatcggagg caagcaggcc ctggaaaccg tgcaaagact gctgcccgtg
     1801 ctgtgccagg cccacggcct cacacccgag caagtggtgg ccatcgctag caacaacggc
     1861 ggcaagcagg ctctcgagac agtccaagct ctgctgccag tgctgtgcca agctcacggc
     1921 ctgacacccg agcaggtggt cgccatcgcc tcccacgatg gcggcaaaca ggctctggaa
     1981 acagtgcagg ccctgctgcc tgtgctgtgc caggcccacg gcctgacccc tgagcaggtg
     2041 gtggccatcg ccagcaatgg cggaggcaag caggccctgg aaaccgtgca ggctctgctg
     2101 cctgtgctgt gtcaagccca tggcctgacc cctgagcagg tggtggctat cgcctctaat
     2161 ggcggaggca agcaggccct ggaaaccgtg cagagactgc tgcctgtgct gtgccaggcc
     2221 catggactga cccctgagca ggttgttgca atcgccagcc acgacggcgg caaacaagcc
     2281 ctggaaaccg tgcaggccct gctgcccgtg ctgtgccagg cccacggact gaccccacag
     2341 caggttgtgg ccatcgccag caacatcggc ggcaagcagg ccctggaaac agtgcagcgg
     2401 ctgctgcctg tgctgtgtca ggcccacggc ctgacccctc aacaggtggt ggccatcgcc
     2461 agccacgacg gcggcaagca ggctcttgag accgtgcaga gactgctgcc cgtgctgtgc
     2521 caggcccacg gactgacacc tcagcaggtg gtggccatcg cctcccacga cggcgggaga
     2581 cctgccctcg aaagcattgt ggctcagctg agccgtccag acccggcgct ggcggctctg
     2641 accaacgatc acctggtggc gctggcttgc ctgggcggtc gtccggccct ggatgccgtg
     2701 aagaaaggcc tgggtggatc cggcagctac gccctgggtc cgtatcagat tagcgccccg
     2761 cagctgccag cttacaatgg tcagaccgtg ggtaccttct actatgtgaa cgacgcgggc
     2821 ggtctggaga gcaaggtgtt tagcagcggc ggtccaaccc cgtacccaaa ctatgccaat
     2881 gccggtcatg tggagggtca gagcgccctg ttcatgcgtg ataacggcat cagcgagggt
     2941 ctggtgttcc acaacaaccc ggaaggcacc tgcggttttt gcgtgaacat gaccgagacc
     3001 ctgctgccgg aaaacgcgaa aatgaccgtg gtgccgccgg aaggttctgg cggctcaact
     3061 aatctgagcg acatcattga gaaggagact gggaaacagc tggtcattca ggagtccatc
     3121 ctgatgctgc ctgaggaggt ggaggaagtg atcggcaaca agccagagtc tgacatcctg
     3181 gtgcacaccg cctacgacga gtccacagat gagaatgtga tgctgctgac ctctgacgcc
     3241 cccgagtata agccttgggc cctggtcatc caggattcta acggcgagaa taagatcaag
     3301 atgctgtgat aaggggtctt tgtcctctgt actgtctctc tccttgcccc taacccaaaa
     3361 agcttcattt ttctgtgtag gctgcacaag agccttgatt gaagatatat tctttctgaa
     3421 cagtatttaa ggtttccaat aaaatgtaca cccctcagaa cttctgaggc ggaaagaacc
     3481 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt
     3541 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     3601 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     3661 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     3721 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     3781 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     3841 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     3901 aatgagctga tttaacaaaa atttaacgcg aattaatttt gtggaatgtg tgtcagttag
     3961 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     4021 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     4081 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     4141 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     4201 aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     4261 gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc
     4321 acgtgttgac aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag
     4381 gaactaaacc atggccaagc ctttgtctca agaagaatcc accctcattg aaagagcaac
     4441 ggctacaatc aacagcatcc ccatctctga agactacagc gtcgccagcg cagctctctc
     4501 tagcgacggc cgcatcttca ctggtgtcaa tgtatatcat tttactgggg gaccttgtgc
     4561 agaactcgtg gtgctgggca ctgctgctgc tgcggcagct ggcaacctga cttgtatcgt
     4621 cgcgatcgga aatgagaaca ggggcatctt gagcccctgc ggacggtgcc gacaggtgct
     4681 tctcgatctg catcctggga tcaaagccat agtgaaggac agtgatggac agccgacggc
     4741 agttgggatt cgtgaattgc tgccctctgg ttatgtgtgg gagggctaag cacttcgtgg
     4801 ccgaggagca ggactgacac gtgctacgag atttcgattc caccgccgcc ttctatgaaa
     4861 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     4921 tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat
     4981 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     5041 gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga
     5101 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     5161 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     5221 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     5281 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     5341 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     5401 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     5461 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     5521 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     5581 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     5641 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     5701 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     5761 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     5821 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     5881 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     5941 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     6001 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     6061 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     6121 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     6181 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     6241 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     6301 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     6361 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     6421 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     6481 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     6541 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     6601 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     6661 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     6721 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     6781 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     6841 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     6901 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     6961 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     7021 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     7081 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     7141 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     7201 tatttgaatg tatttagaaa aataaacaaa taggggtt
//
