LOCUS       sequence-416871-        7304 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To install m.G3635 mutation on mtDNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7304)
  AUTHORS   Sun H, Wang Z, Shen L, Feng Y, Han L, Qian X, Meng R, Ji K, Liang D,
            Zhou F, Lou X, Zhang J, Shen B
  TITLE     Developing mitochondrial base editors with diverse context 
            compatibility and high fidelity via saturated spacer library.
  JOURNAL   Nat Commun. 2023 Oct 19;14(1):6625. doi: 10.1038/s41467-023-42359-3.
  PUBMED    37857619
REFERENCE   2  (bases 1 to 7304)
  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"; journalName: "Nat 
            Commun."; date: "2023-10-19"; volume: "14(1):6625. doi"; pages: " 
            10.1038/s41467-023-42359-3"
COMMENT     Sequence Label: pGL3-MTS-G3635A-L17-Q2L7-T2176C-UGI-Puro
FEATURES             Location/Qualifiers
     source          1..7304
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        8..387
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        388..591
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     542..562
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     636..655
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        636..654
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             863..886
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3212..3466
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKMLA"
     primer_bind     complement(3499..3516)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3505..3729
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        3736..4246
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     4165..4183
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             4268..4867
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4268..4287)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4764..4784
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5014..5135
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5025..5044)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5079..5098
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(5383..5400)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5554..6142)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5634..5653)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6313..7104,7105..7173))
                     /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     6936..6955
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7174..7278)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 acgcgttgac attgattatt gactagttat taatagtaat caattacggg gtcattagtt
       61 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
      121 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
      181 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
      241 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
      301 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
      361 tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt
      421 ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt
      481 ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg
      541 acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc tctctggcta
      601 actagagaac ccactgctta ctggcttatc gaaattaata cgactcacta tagggagaca
      661 agctggctag accatgttgg ggtttgtggg tcgggtggcc gctgctccgg cctccggggc
      721 cttgcggaga ctcacccctt cagcgtcgct gcccccagct cagctcttac tgcgggccgc
      781 tccgacggcg gtccatcctg tcagggacta tgcggcgcaa acatctgaga gcggcggagg
      841 cgggagcccc ggcgcggccg ctgactacaa ggatgacgac gataaaggta gcgtggatct
      901 acgcacgctc ggctacagcc agcagcaaca ggagaagatc aaaccgaagg ttcgttcgac
      961 agtggcgcag caccacgagg cactggtcgg ccatgggttt acacacgcgc acatcgttgc
     1021 gctcagccaa cacccggcag cgttagggac cgtcgctgtc aagtatcagg acatgatcgc
     1081 agcgttgcca gaggcgacac acgaagcgat cgttggcgtc ggcaaacggg gcgccggcgc
     1141 acgcgctctg gaggccttgc tcacggtggc gggagagttg agaggtccac cgttacagtt
     1201 ggacacaggc caacttctca agattgcaaa acgtggcggc gtgaccgcag tggaggcagt
     1261 gcatgcatgg cgcaatgcac tgacgggtgc ccccctgaac ctgaccccgg agcaggtggt
     1321 ggccatcgct agtcatgacg gtggcaaaca ggctcttgaa accgtccaac gccttctacc
     1381 agttctctgt caagcccacg gactaacccc agcgcaagtt gtagcgattg ctagtcatga
     1441 cggtggcaaa caggctcttg aaaccgtcca acgccttcta ccagttctct gtcaagccca
     1501 cggactaacc ccagcgcagg ttgtagcgat tgctagtaat gggggtggca aacaggctct
     1561 tgagactgtt cagcgccttc taccagttct ctgtcaagcc cacggcctga cgcccgagca
     1621 agttgtagcg attgctagta atattggtgg caaacaggct cttgagacgg ttcagcgcct
     1681 ccttccagtt ctctgtcaag cccacggact caccccagat caagtggtag cgattgctag
     1741 taatgggggt ggcaaacagg ctcttgagac tgttcagcgc cttctaccag ttctctgtca
     1801 agcccacggc ctgacgcccg agcaagttgt agcgattgct agtaatgggg gtggcaaaca
     1861 ggctcttgag actgttcagc gccttctacc agttctctgt caagcccacg gcctgacgcc
     1921 cgagcaagtt gtagcgattg ctagtaatgg gggtggcaaa caggctcttg agactgttca
     1981 gcgccttcta ccagttctct gtcaagccca cggcctgacg cccgagcaag ttgtagcgat
     2041 tgctagtaat attggtggca aacaggctct tgagacggtt cagcgcctcc ttccagttct
     2101 ctgtcaagcc cacggactca ccccagatca agttgtagca attgctagta atgggggtgg
     2161 caaacaggct cttgagactg ttcagcgcct tctaccagtt ctctgtcaag cccacggcct
     2221 gacgcccgag caagttgtag cgattgctag taatgggggt ggcaaacagg ctcttgagac
     2281 tgttcagcgc cttctaccag ttctctgtca agcccacggc ctgacgcccg agcaagttgt
     2341 agcgattgct agtcatgacg gtggcaaaca ggctcttgaa accgtccaac gccttctacc
     2401 agttctctgt caagcccacg gactaacccc agcgcaagtt gtagcgattg ctagtaatgg
     2461 gggtggcaaa caggctcttg agactgttca gcgccttcta ccagttctct gtcaagccca
     2521 cggcctgacg cccgagcaag ttgtagcgat agccagtaat attggtggca aacaggctct
     2581 tgagacggtt cagcgcctcc ttccagttct ctgtcaagcc cacggactca ccccagatca
     2641 agttgtagcg attgctagta acaatggtgg caaacaggct cttgaaaccg tacagcgcct
     2701 actgccagtt ctctgtcaag cccacggtct gactccggag caagttgtag cgattgcaag
     2761 tcatgacggt ggcaaacagg ctcttgaaac cgtccaacgc cttctaccag ttctctgtca
     2821 agcccacgga ctaaccccag cgcaagttgt agcgattgct agtcatgacg gtggcaaaca
     2881 ggctcttgaa accgtccaac gccttctacc agttctctgt caagcccacg gactaacccc
     2941 agcgcaagtt gtagcgattg cttccaatat tggcggcagg ccggcgctgg agagcattgt
     3001 tgcccagtta tctcgccctg atccggcgtt ggccgcgttg accaacgacc acctcgtcgc
     3061 gctagcctgc ctcggcggac gtcctgcgct ggatgcagtg aaaaagggat tgggaggctc
     3121 caccgtgcca ccaaccgcaa ggtggagcaa cggcaagacc ttcgtgggca acgagaacga
     3181 tccaaagcca tggccaaggt ctggaggcag caccaacctg tctgacatca tcgagaagga
     3241 gacaggcaag cagctggtca tccaggagag catcctgatg ctgcccgaag aagtcgaaga
     3301 agtgatcgga aacaagcctg agagcgatat cctggtccat accgcctacg acgagagtac
     3361 cgacgaaaat gtgatgctgc tgacatccga cgccccagag tataagccct gggctctggt
     3421 catccaggat tccaacggag agaacaaaat caaaatgctg gcttaagtct agagggcccg
     3481 tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc
     3541 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
     3601 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
     3661 ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg
     3721 gctctatggc tcgagggggt tggggttgcg ccttttccaa ggcagccctg ggtttgcgca
     3781 gggacgcggc tgctctgggc gtggttccgg gaaacgcagc ggcgccgacc ctgggactcg
     3841 cacattcttc acgtccgttc gcagcgtcac ccggatcttc gccgctaccc ttgtgggccc
     3901 cccggcgacg cttcctgctc cgcccctaag tcgggaaggt tccttgcggt tcgcggcgtg
     3961 ccggacgtga caaacggaag ccgcacgtct cactagtacc ctcgcagacg gacagcgcca
     4021 gggagcaatg gcagcgcgcc gaccgcgatg ggctgtggcc aatagcggct gctcagcagg
     4081 gcgcgccgag agcagcggcc gggaaggggc ggtgcgggag gcggggtgtg gggcggtagt
     4141 gtgggccctg ttcctgcccg cgcggtgttc cgcattctgc aagcctccgg agcgcacgtc
     4201 ggcagtcggc tccctcgttg accgaatcac cgacctctct ccccaggggg atccaccgga
     4261 gcttaccatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag
     4321 ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga
     4381 tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg
     4441 gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac
     4501 gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt
     4561 gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc
     4621 caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg
     4681 tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc
     4741 cttcctggaa acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt
     4801 caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg
     4861 tgcctgacgc ccgccccacg acccgcagcg cccgaccgaa aggagcgcac gaccccatgc
     4921 atcggtacct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttctaga
     4981 gtcggggcgg ccggccgctt cgagcagaca tgataagata cattgatgag tttggacaaa
     5041 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     5101 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     5161 tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac ctctacaaat
     5221 gtggtaaaat cgataaggat ccgtcgaccg atgcccttga gagccttcaa cccagtcagc
     5281 tccttccggt gggcgcgggg catgactatc gtcgccgcac ttatgactgt cttctttatc
     5341 atgcaactcg taggacaggt gccggcagcg ctcttccgct tcctcgctca ctgactcgct
     5401 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     5461 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     5521 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     5581 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     5641 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     5701 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     5761 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     5821 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     5881 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     5941 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     6001 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     6061 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     6121 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     6181 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     6241 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     6301 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     6361 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6421 accatctggc cccagtgctg caatgatacc gcgggaccca cgctcaccgg ctccagattt
     6481 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6541 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     6601 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     6661 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     6721 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     6781 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     6841 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     6901 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6961 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     7021 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     7081 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     7141 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     7201 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     7261 acaaataggg gttccgcgca catttccccg aaaagtgcca cctg
//
