LOCUS       sequence-416799-        7297 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 7297)
  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 7297)
  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-FZY2-S100C-UGI-Puro
FEATURES             Location/Qualifiers
     source          1..7297
                     /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             3209..3460
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     primer_bind     complement(3492..3509)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3498..3722
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        3729..4239
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     4158..4176
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             4261..4860
                     /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(4261..4280)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4757..4777
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5007..5128
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5018..5037)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5072..5091
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(5376..5393)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5547..6135)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5627..5646)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6306..7097,7098..7166))
                     /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     6929..6948
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7167..7271)
                     /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 cagggcaatc gattacgtca agacctacac cggcaccagc aacgatccaa agatcagccc
     3181 aaggtacaag ggcaactctg gaggcagcac caacctgtct gacatcatcg agaaggagac
     3241 aggcaagcag ctggtcatcc aggagagcat cctgatgctg cccgaagaag tcgaagaagt
     3301 gatcggaaac aagcctgaga gcgatatcct ggtccatacc gcctacgacg agagtaccga
     3361 cgaaaatgtg atgctgctga catccgacgc cccagagtat aagccctggg ctctggtcat
     3421 ccaggattcc aacggagaga acaaaatcaa aatgctgtaa tctagagggc ccgtttaaac
     3481 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     3541 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     3601 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     3661 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     3721 ggctcgaggg ggttggggtt gcgccttttc caaggcagcc ctgggtttgc gcagggacgc
     3781 ggctgctctg ggcgtggttc cgggaaacgc agcggcgccg accctgggac tcgcacattc
     3841 ttcacgtccg ttcgcagcgt cacccggatc ttcgccgcta cccttgtggg ccccccggcg
     3901 acgcttcctg ctccgcccct aagtcgggaa ggttccttgc ggttcgcggc gtgccggacg
     3961 tgacaaacgg aagccgcacg tctcactagt accctcgcag acggacagcg ccagggagca
     4021 atggcagcgc gccgaccgcg atgggctgtg gccaatagcg gctgctcagc agggcgcgcc
     4081 gagagcagcg gccgggaagg ggcggtgcgg gaggcggggt gtggggcggt agtgtgggcc
     4141 ctgttcctgc ccgcgcggtg ttccgcattc tgcaagcctc cggagcgcac gtcggcagtc
     4201 ggctccctcg ttgaccgaat caccgacctc tctccccagg gggatccacc ggagcttacc
     4261 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta
     4321 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac
     4381 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac
     4441 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag
     4501 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt
     4561 tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag
     4621 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc
     4681 agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg
     4741 gaaacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc
     4801 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga
     4861 cgcccgcccc acgacccgca gcgcccgacc gaaaggagcg cacgacccca tgcatcggta
     4921 cctttaagac caatgactta caaggcagct gtagatctta gccactttct agagtcgggg
     4981 cggccggccg cttcgagcag acatgataag atacattgat gagtttggac aaaccacaac
     5041 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     5101 aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca
     5161 ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtaa
     5221 aatcgataag gatccgtcga ccgatgccct tgagagcctt caacccagtc agctccttcc
     5281 ggtgggcgcg gggcatgact atcgtcgccg cacttatgac tgtcttcttt atcatgcaac
     5341 tcgtaggaca ggtgccggca gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     5401 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5461 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5521 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5581 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     5641 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     5701 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     5761 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     5821 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     5881 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     5941 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     6001 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     6061 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     6121 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     6181 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     6241 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     6301 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     6361 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     6421 ggccccagtg ctgcaatgat accgcgggac ccacgctcac cggctccaga tttatcagca
     6481 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     6541 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     6601 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     6661 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     6721 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     6781 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     6841 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     6901 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     6961 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     7021 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     7081 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     7141 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     7201 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     7261 ggggttccgc gcacatttcc ccgaaaagtg ccacctg
//
