LOCUS       sequence-355098-        8012 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression  of base editor in mammalian cells and for cell sorting .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8012)
  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 8012)
  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-DddA11-mCherry
FEATURES             Location/Qualifiers
     source          1..8012
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(109..128)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        300..679
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        680..883
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     834..854
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     928..947
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        928..946
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             969..1046
                     /codon_start=1
                     /product="mitochondrial presequence of human cytochrome c 
                     oxidase subunit VIII (Rizutto et al., 1989)"
                     /label=COX8 presequence
                     /translation="MASVLTPLLLRGLTGSARRLPVPRAK"
     CDS             1059..1124
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3087..3335
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             3345..3401
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     rep_origin      4309..4737
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4396..4415)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4606..4627
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4746..4766)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4751..5080
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4931..5066
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4993..5012
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5128..5175
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5194..5592
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    5750..5871
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5787..5806)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5841..5860
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5920..5936)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5920..5936)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5933..5955)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5944..5960
                     /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(5968..5974,5975..5992,5993..5998))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6013..6034
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6151..6168)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6322..6910)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6402..6421)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7081..7872,7873..7941))
                     /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     7704..7723
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7942..8012,1..34))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
       61 acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta caatctgctc
      121 tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta
      181 gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa
      241 tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg
      301 acattgatta ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc
      361 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
      421 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      481 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      541 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      601 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      661 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
      721 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
      781 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
      841 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc taactagaga
      901 acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg
      961 ctagcaccat ggcctccgtt ctgaccccgc tgctgctgcg tggcctgacc ggctccgctc
     1021 gtcgtctgcc agttccgcgt gcgaaaatcc attccctgga ctacaaagac catgacggtg
     1081 attataaaga tcatgacatc gattacaagg atgacgatga caagatggac atcgcggatc
     1141 tgcgtaccct gggttacagc cagcagcagc aggagaagat caagccgaag gtgcgcagca
     1201 ccgtggctca gcaccacgaa gccctggtgg gccacggttt cacccacgct cacatcgtgg
     1261 ccctgagcca gcacccagcc gcgctgggca ccgtggccgt gaaatatcag gacatgattg
     1321 ctgccctgcc agaggccacc catgaagcta ttgtgggcgt gggcaagcag tggagcggtg
     1381 ctcgtgcgct ggaggcgctg ctgaccgtgg ctggtgaact gcgtggtccg ccgctgcagc
     1441 tggataccgg tcagctgctg aaaattgcca aacgtggcgg tgtgaccgcg gtggaagccg
     1501 tgcatgcttg gcgtaatgct ctgaccggtg cgccgctgaa cttaacgccg gaacaagttg
     1561 ttgcaattgc cagcaacaac ggcggcaagc aagctctgga aaccgtgcag gccctgcttc
     1621 cagtcctgtg tcaggcccac ggcctgaccc ctcagcaggt ggtcgccatc gcctctaaca
     1681 tcggcggaaa gcaggctctg gagaccgtgc agcggctgct gcccgtgctg tgccaggccc
     1741 acggcctgac acctcagcag gtggtggcca ttgctagcaa catcggaggc aagcaggccc
     1801 tggaaaccgt gcaaagactg ctgcccgtgc tgtgccaggc ccacggcctc acacccgagc
     1861 aagtggtggc catcgctagc aacaacggcg gcaagcaggc tctcgagaca gtccaagctc
     1921 tgctgccagt gctgtgccaa gctcacggcc tgacacccga gcaggtggtc gccatcgcct
     1981 cccacgatgg cggcaaacag gctctggaaa cagtgcaggc cctgctgcct gtgctgtgcc
     2041 aggcccacgg cctgacccct gagcaggtgg tggccatcgc cagcaatggc ggaggcaagc
     2101 aggccctgga aaccgtgcag gctctgctgc ctgtgctgtg tcaagcccat ggcctgaccc
     2161 ctgagcaggt ggtggctatc gcctctaatg gcggaggcaa gcaggccctg gaaaccgtgc
     2221 agagactgct gcctgtgctg tgccaggccc atggactgac ccctgagcag gttgttgcaa
     2281 tcgccagcca cgacggcggc aaacaagccc tggaaaccgt gcaggccctg ctgcccgtgc
     2341 tgtgccaggc ccacggactg accccacagc aggttgtggc catcgccagc aacatcggcg
     2401 gcaagcaggc cctggaaaca gtgcagcggc tgctgcctgt gctgtgtcag gcccacggcc
     2461 tgacccctca acaggtggtg gccatcgcca gccacgacgg cggcaagcag gctcttgaga
     2521 ccgtgcagag actgctgccc gtgctgtgcc aggcccacgg actgacacct cagcaggtgg
     2581 tggccatcgc ctcccacgac ggcgggagac ctgccctcga aagcattgtg gctcagctga
     2641 gccgtccaga cccggcgctg gcggctctga ccaacgatca cctggtggcg ctggcttgcc
     2701 tgggcggtcg tccggccctg gatgccgtga agaaaggcct gggtggatcc ggcagctacg
     2761 ccctgggtcc gtatcagatt agcgccccgc agctgccagc ttacaatggt cagaccgtgg
     2821 gtaccttcta ctatgtgaac gacgcgggcg gtctggagag caaggtgttt atcagcggcg
     2881 gtccaacccc gtacccaaac tatgtcagtg ccggtcatgt ggagggtcag agcgccctgt
     2941 tcatgcgtga taacggcatc agcgagggtc tggtgttcca caacaacccg aaaggcacct
     3001 gcggtttttg cgtgaacatg atcgagaccc tgctgccgga aaacgcgaaa atgaccgtgg
     3061 tgccgccgga aggttctggc ggctcaacta atctgagcga catcattgag aaggagactg
     3121 ggaaacagct ggtcattcag gagtccatcc tgatgctgcc tgaggaggtg gaggaagtga
     3181 tcggcaacaa gccagagtct gacatcctgg tgcacaccgc ctacgacgag tccacagatg
     3241 agaatgtgat gctgctgacc tctgacgccc ccgagtataa gccttgggcc ctggtcatcc
     3301 aggattctaa cggcgagaat aagatcaaga tgctgggaag cggagctact aacttcagcc
     3361 tgctgaagca ggctggagac gtggaggaga accctggacc tatggtgtcc aagggcgagg
     3421 aagataacat ggccatcatc aaggagttca tgaggtttaa ggtccacatg gagggttcag
     3481 tcaatggcca cgagttcgag attgaaggcg agggcgaggg ccgcccctac gaagggacac
     3541 agacggcgaa attgaaggtg accaaaggcg ggccattgcc cttcgcatgg gacatcttgt
     3601 cccctcagtt tatgtatggc agcaaggcct atgttaagca ccccgctgat atcccggact
     3661 acttgaagct gtcctttcca gaggggttta aatgggagcg cgttatgaat ttcgaagacg
     3721 gaggagtggt tacggtgacg caggactcat ccctgcagga cggagaattt atatataagg
     3781 ttaagttgag aggcacaaac ttcccaagcg acggccctgt gatgcagaag aaaacaatgg
     3841 ggtgggaagc ttccagcgag cgcatgtacc ccgaagatgg cgccctcaag ggcgagataa
     3901 agcaaaggct gaaacttaag gacggcggtc attacgacgc ggaggtcaag acaacttaca
     3961 aggctaaaaa acccgttcag ttgcctgggg cttacaatgt taatatcaaa cttgacatca
     4021 caagccacaa tgaagactat acgatcgtgg agcagtatga acgagcggaa ggcaggcact
     4081 caacgggggg gatggacgag ctttacaagt gataaggggt ctttgtcctc tgtactgtct
     4141 ctctccttgc ccctaaccca aaaagcttca tttttctgtg taggctgcac aagagccttg
     4201 attgaagata tattctttct gaacagtatt taaggtttcc aataaaatgt acacccctca
     4261 gaacttctga ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta
     4321 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     4381 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     4441 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     4501 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     4561 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     4621 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     4681 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat
     4741 tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
     4801 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
     4861 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
     4921 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     4981 actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa
     5041 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat
     5101 atccattttc ggatctgatc agcacgtgtt gacaattaat catcggcata gtatatcggc
     5161 atagtataat acgacaaggt gaggaactaa accatggcca agcctttgtc tcaagaagaa
     5221 tccaccctca ttgaaagagc aacggctaca atcaacagca tccccatctc tgaagactac
     5281 agcgtcgcca gcgcagctct ctctagcgac ggccgcatct tcactggtgt caatgtatat
     5341 cattttactg ggggaccttg tgcagaactc gtggtgctgg gcactgctgc tgctgcggca
     5401 gctggcaacc tgacttgtat cgtcgcgatc ggaaatgaga acaggggcat cttgagcccc
     5461 tgcggacggt gccgacaggt gcttctcgat ctgcatcctg ggatcaaagc catagtgaag
     5521 gacagtgatg gacagccgac ggcagttggg attcgtgaat tgctgccctc tggttatgtg
     5581 tgggagggct aagcacttcg tggccgagga gcaggactga cacgtgctac gagatttcga
     5641 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     5701 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
     5761 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     5821 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     5881 tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     5941 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     6001 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     6061 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     6121 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     6181 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     6241 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     6301 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     6361 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     6421 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     6481 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     6541 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     6601 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     6661 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     6721 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     6781 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     6841 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     6901 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     6961 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     7021 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     7081 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     7141 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     7201 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     7261 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     7321 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     7381 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     7441 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     7501 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     7561 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     7621 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     7681 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     7741 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     7801 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     7861 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     7921 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     7981 ttattgtctc atgagcggat acatatttga at
//
