LOCUS       sequence-416899-        7546 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 7546)
  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 7546)
  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-T2176N-UGI-Puro
FEATURES             Location/Qualifiers
     source          1..7546
                     /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             3458..3709
                     /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(3741..3758)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3747..3971
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        3978..4488
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     4407..4425
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             4510..5109
                     /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(4510..4529)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5006..5026
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5256..5377
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5267..5286)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5321..5340
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(5625..5642)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5796..6384)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5876..5895)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6555..7346,7347..7415))
                     /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     7178..7197
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7416..7520)
                     /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 cgggaaggtg ggcaagttga ggtttgcacc aaaggtggag agcgcagaga gcatgttgag
     3181 gagcttgagc caggagggca agaccgcagg cgtgttggat atcaacggcg agttgatccc
     3241 attggtgagc ggcaccagca gcttgaagaa ctacgcagca agcggccatg tggagggcca
     3301 agcagcattg atcatgaggg agaggggcgt ggcaagcgca aggcttatca tcgataaccc
     3361 aagcggcatc tgcggctact gcaggagcca ggtgccaacc ttgttgccag caggcgcaac
     3421 cttggaggtg accaccccaa ggggctctgg aggcagcacc aacctgtctg acatcatcga
     3481 gaaggagaca ggcaagcagc tggtcatcca ggagagcatc ctgatgctgc ccgaagaagt
     3541 cgaagaagtg atcggaaaca agcctgagag cgatatcctg gtccataccg cctacgacga
     3601 gagtaccgac gaaaatgtga tgctgctgac atccgacgcc ccagagtata agccctgggc
     3661 tctggtcatc caggattcca acggagagaa caaaatcaaa atgctgtaat ctagagggcc
     3721 cgtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     3781 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     3841 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     3901 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
     3961 gggctctatg gctcgagggg gttggggttg cgccttttcc aaggcagccc tgggtttgcg
     4021 cagggacgcg gctgctctgg gcgtggttcc gggaaacgca gcggcgccga ccctgggact
     4081 cgcacattct tcacgtccgt tcgcagcgtc acccggatct tcgccgctac ccttgtgggc
     4141 cccccggcga cgcttcctgc tccgccccta agtcgggaag gttccttgcg gttcgcggcg
     4201 tgccggacgt gacaaacgga agccgcacgt ctcactagta ccctcgcaga cggacagcgc
     4261 cagggagcaa tggcagcgcg ccgaccgcga tgggctgtgg ccaatagcgg ctgctcagca
     4321 gggcgcgccg agagcagcgg ccgggaaggg gcggtgcggg aggcggggtg tggggcggta
     4381 gtgtgggccc tgttcctgcc cgcgcggtgt tccgcattct gcaagcctcc ggagcgcacg
     4441 tcggcagtcg gctccctcgt tgaccgaatc accgacctct ctccccaggg ggatccaccg
     4501 gagcttacca tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc
     4561 agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc
     4621 gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc
     4681 gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc
     4741 acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag
     4801 ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg
     4861 cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag
     4921 ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc
     4981 gccttcctgg aaacctccgc gccccgcaac ctccccttct acgagcggct cggcttcacc
     5041 gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc
     5101 ggtgcctgac gcccgcccca cgacccgcag cgcccgaccg aaaggagcgc acgaccccat
     5161 gcatcggtac ctttaagacc aatgacttac aaggcagctg tagatcttag ccactttcta
     5221 gagtcggggc ggccggccgc ttcgagcaga catgataaga tacattgatg agtttggaca
     5281 aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc
     5341 tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt
     5401 tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa acctctacaa
     5461 atgtggtaaa atcgataagg atccgtcgac cgatgccctt gagagccttc aacccagtca
     5521 gctccttccg gtgggcgcgg ggcatgacta tcgtcgccgc acttatgact gtcttcttta
     5581 tcatgcaact cgtaggacag gtgccggcag cgctcttccg cttcctcgct cactgactcg
     5641 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     5701 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     5761 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     5821 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     5881 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     5941 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     6001 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     6061 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     6121 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     6181 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     6241 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     6301 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     6361 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     6421 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     6481 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     6541 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     6601 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     6661 ttaccatctg gccccagtgc tgcaatgata ccgcgggacc cacgctcacc ggctccagat
     6721 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     6781 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     6841 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     6901 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     6961 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     7021 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     7081 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     7141 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     7201 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     7261 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     7321 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     7381 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     7441 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     7501 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctg
//
