LOCUS       sequence-352080-        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-DddA6
FEATURES             Location/Qualifiers
     source          1..7238
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(133..152)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        324..703
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        704..907
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     858..878
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     952..971
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        952..970
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             993..1070
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MASVLTPLLLRGLTGSARRLPVPRAK"
     CDS             1083..1148
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3111..3362
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     rep_origin      3559..3987
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3646..3665)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3856..3877
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3996..4016)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4001..4330
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4181..4316
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4243..4262
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4378..4425
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4444..4842
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    5000..5121
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5037..5056)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5091..5110
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5170..5186)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5170..5186)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5183..5205)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5194..5210
                     /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(5218..5224,5225..5242,5243..5248))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5263..5284
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5401..5418)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5572..6160)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5652..5671)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6331..7122,7123..7191))
                     /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     6954..6973
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7192..7238,1..58))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
       61 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg
      121 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc
      181 ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg
      241 cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga
      301 tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat
      361 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
      421 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
      481 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
      541 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
      601 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
      661 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
      721 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
      781 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
      841 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
      901 cagagctctc tggctaacta gagaacccac tgcttactgg cttatcgaaa ttaatacgac
      961 tcactatagg gagacccaag ctggctagca ccatggcctc cgttctgacc ccgctgctgc
     1021 tgcgtggcct gaccggctcc gctcgtcgtc tgccagttcc gcgtgcgaaa atccattccc
     1081 tggactacaa agaccatgac ggtgattata aagatcatga catcgattac aaggatgacg
     1141 atgacaagat ggacatcgcg gatctgcgta ccctgggtta cagccagcag cagcaggaga
     1201 agatcaagcc gaaggtgcgc agcaccgtgg ctcagcacca cgaagccctg gtgggccacg
     1261 gtttcaccca cgctcacatc gtggccctga gccagcaccc agccgcgctg ggcaccgtgg
     1321 ccgtgaaata tcaggacatg attgctgccc tgccagaggc cacccatgaa gctattgtgg
     1381 gcgtgggcaa gcagtggagc ggtgctcgtg cgctggaggc gctgctgacc gtggctggtg
     1441 aactgcgtgg tccgccgctg cagctggata ccggtcagct gctgaaaatt gccaaacgtg
     1501 gcggtgtgac cgcggtggaa gccgtgcatg cttggcgtaa tgctctgacc ggtgcgccgc
     1561 tgaacttaac gccggaacaa gttgttgcaa ttgccagcaa caacggcggc aagcaagctc
     1621 tggaaaccgt gcaggccctg cttccagtcc tgtgtcaggc ccacggcctg acccctcagc
     1681 aggtggtcgc catcgcctct aacatcggcg gaaagcaggc tctggagacc gtgcagcggc
     1741 tgctgcccgt gctgtgccag gcccacggcc tgacacctca gcaggtggtg gccattgcta
     1801 gcaacatcgg aggcaagcag gccctggaaa ccgtgcaaag actgctgccc gtgctgtgcc
     1861 aggcccacgg cctcacaccc gagcaagtgg tggccatcgc tagcaacaac ggcggcaagc
     1921 aggctctcga gacagtccaa gctctgctgc cagtgctgtg ccaagctcac ggcctgacac
     1981 ccgagcaggt ggtcgccatc gcctcccacg atggcggcaa acaggctctg gaaacagtgc
     2041 aggccctgct gcctgtgctg tgccaggccc acggcctgac ccctgagcag gtggtggcca
     2101 tcgccagcaa tggcggaggc aagcaggccc tggaaaccgt gcaggctctg ctgcctgtgc
     2161 tgtgtcaagc ccatggcctg acccctgagc aggtggtggc tatcgcctct aatggcggag
     2221 gcaagcaggc cctggaaacc gtgcagagac tgctgcctgt gctgtgccag gcccatggac
     2281 tgacccctga gcaggttgtt gcaatcgcca gccacgacgg cggcaaacaa gccctggaaa
     2341 ccgtgcaggc cctgctgccc gtgctgtgcc aggcccacgg actgacccca cagcaggttg
     2401 tggccatcgc cagcaacatc ggcggcaagc aggccctgga aacagtgcag cggctgctgc
     2461 ctgtgctgtg tcaggcccac ggcctgaccc ctcaacaggt ggtggccatc gccagccacg
     2521 acggcggcaa gcaggctctt gagaccgtgc agagactgct gcccgtgctg tgccaggccc
     2581 acggactgac acctcagcag gtggtggcca tcgcctccca cgacggcggg agacctgccc
     2641 tcgaaagcat tgtggctcag ctgagccgtc cagacccggc gctggcggct ctgaccaacg
     2701 atcacctggt ggcgctggct tgcctgggcg gtcgtccggc cctggatgcc gtgaagaaag
     2761 gcctgggtgg atccggcagc tacgccctgg gtccgtatca gattagcgcc ccgcagctgc
     2821 cagcttacaa tggtcggacc gtgggtacct tctactatgt gaacgacgcg ggcggtctgg
     2881 agagcaaggt gtttatcagc ggcggtccaa ccccgtaccc aaactatgcc aatgccggtc
     2941 atgtggaggg tcagagcgcc ctgttcatgc gtgataacgg catcagcgag ggtctggtgt
     3001 tccacaacaa cccggaaggc acctgcggtt tttgcgtgaa catgatcgag accctgctgc
     3061 cggaaaacgc gaaaatgacc gtggtgccgc cggaaggttc tggcggctca actaatctga
     3121 gcgacatcat tgagaaggag actgggaaac agctggtcat tcaggagtcc atcctgatgc
     3181 tgcctgagga ggtggaggaa gtgatcggca acaagccaga gtctgacatc ctggtgcaca
     3241 ccgcctacga cgagtccaca gatgagaatg tgatgctgct gacctctgac gcccccgagt
     3301 ataagccttg ggccctggtc atccaggatt ctaacggcga gaataagatc aagatgctgt
     3361 gataaggggt ctttgtcctc tgtactgtct ctctccttgc ccctaaccca aaaagcttca
     3421 tttttctgtg taggctgcac aagagccttg attgaagata tattctttct gaacagtatt
     3481 taaggtttcc aataaaatgt acacccctca gaacttctga ggcggaaaga accagctggg
     3541 gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     3601 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     3661 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     3721 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     3781 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     3841 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     3901 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     3961 tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg
     4021 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     4081 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     4141 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     4201 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     4261 ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     4321 cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc agcacgtgtt
     4381 gacaattaat catcggcata gtatatcggc atagtataat acgacaaggt gaggaactaa
     4441 accatggcca agcctttgtc tcaagaagaa tccaccctca ttgaaagagc aacggctaca
     4501 atcaacagca tccccatctc tgaagactac agcgtcgcca gcgcagctct ctctagcgac
     4561 ggccgcatct tcactggtgt caatgtatat cattttactg ggggaccttg tgcagaactc
     4621 gtggtgctgg gcactgctgc tgctgcggca gctggcaacc tgacttgtat cgtcgcgatc
     4681 ggaaatgaga acaggggcat cttgagcccc tgcggacggt gccgacaggt gcttctcgat
     4741 ctgcatcctg ggatcaaagc catagtgaag gacagtgatg gacagccgac ggcagttggg
     4801 attcgtgaat tgctgccctc tggttatgtg tgggagggct aagcacttcg tggccgagga
     4861 gcaggactga cacgtgctac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     4921 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     4981 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     5041 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     5101 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     5161 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     5221 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     5281 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     5341 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     5401 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     5461 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     5521 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     5581 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     5641 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     5701 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     5761 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     5821 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     5881 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     5941 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     6001 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     6061 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     6121 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     6181 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     6241 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     6301 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     6361 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     6421 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     6481 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     6541 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     6601 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     6661 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     6721 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     6781 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     6841 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     6901 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     6961 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     7021 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     7081 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     7141 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     7201 ttttcaatat tattgaagca tttatcaggg ttattgtc
//
