LOCUS       sequence-405608-        6318 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A backbone containing a RsDddA fragment C for constructing 
            mitochondria-targeted RsDdCBE expression plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6318)
  AUTHORS   Cheng K, Li C, Jin J, Qian X, Guo J, Shen L, Dai Y, Zhang X, Li Z, 
            Guan Y, Zhou F, Tang J, Zhang J, Shen B, Lou X
  TITLE     Engineering RsDddA as mitochondrial base editor with wide target 
            compatibility and enhanced activity.
  JOURNAL   Mol Ther Nucleic Acids. 2023 Sep 9;34:102028. doi: 
            10.1016/j.omtn.2023.09.005. eCollection 2023 Dec 12.
  PUBMED    37744175
REFERENCE   2  (bases 1 to 6318)
  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: "Mol Ther 
            Nucleic Acids."; date: "2023-09-9"; volume: "34:102028. doi"; pages:
            " 10.1016/j.omtn.2023.09.005. eCollection 2023 Dec 12"
COMMENT     Sequence Label: pGL3-MTS-CCDB-N136-RsDddA-C-UGI-PURO
FEATURES             Location/Qualifiers
     source          1..6318
                     /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             1645..1950
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     1829..1848
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             2226..2480
                     /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(2513..2530)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2519..2743
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        2750..3260
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3179..3197
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3282..3881
                     /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(3282..3301)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3778..3798
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4028..4149
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4039..4058)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4093..4112
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4397..4414)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4568..5156)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4648..4667)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5327..6118,6119..6187))
                     /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     5950..5969
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6188..6292)
                     /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 ggcaaacagt ggtccggcgc
     1141 acgcgctctg gaggccttgc tcacggtggc gggagagttg agaggtccac cgttacagtt
     1201 ggacacaggc caacttctca agattgcaaa acgtggcggc gtgaccgcag tggaggcagt
     1261 gcatgcatgg cgcaatgcac tgacgggtgc ccccctgaac ctgaccccag agaccggctt
     1321 actaaaagcc agataacagt atgcgtattt gcgcgctgat ttttgcggta taagaatata
     1381 tactgatatg tatacccgaa gtatgtcaaa aagaggtatg ctatgaagca gcgtattaca
     1441 gtgacagttg acagcgacag ctatcagttg ctcaaggcat atatgatgtc aatatctccg
     1501 gtctggtaag cacaaccatg cagaatgaag cccgtcgtct gcgtgccgaa cgctggaaag
     1561 cggaaaatca ggaagggatg gctgaggtcg cccggtttat tgaaatgaac ggctcttttg
     1621 ctgacgagaa caggggctgg tgaaatgcag tttaaggttt acacctataa aagagagagc
     1681 cgttatcgtc tgtttgtgga tgtacagagt gatattattg acacgcccgg gcgacggatg
     1741 gtgatccccc tggccagtgc acgtctgctg tcagataaag tctcccgtga actttacccg
     1801 gtggtgcata tcggggatga aagctggcgc atgatgacca ccgatatggc cagtgtgccg
     1861 gtgtccgtta tcggggaaga agtggctgat ctcagccacc gcgaaaatga catcaaaaac
     1921 gccattaacc tgatgttctg gggaatataa atgtcaggct cccttataca cagccagggt
     1981 ctctcgcgtt gaccaacgac cacctcgtcg ccttggcctg cctcggcgga cgtcctgcgc
     2041 tggatgcagt gaaaaaggga ttgggaggct ccgctagcaa gtgcagactg ctggtgcaca
     2101 acgccaactg caaccaggag aagcccgtgc tgcccaagta cgacggcaag accaccgagg
     2161 gcgtgatggt gacccccgac ggcaagcaga tcagcttcaa gtccggcaac gcatcaggcg
     2221 ggagcaccaa cctgtctgac atcatcgaga aggagacagg caagcagctg gtcatccagg
     2281 agagcatcct gatgctgccc gaagaagtcg aagaagtgat cggaaacaag cctgagagcg
     2341 atatcctggt ccataccgcc tacgacgaga gtaccgacga aaatgtgatg ctgctgacat
     2401 ccgacgcccc agagtataag ccctgggctc tggtcatcca ggattccaac ggagagaaca
     2461 aaatcaaaat gctggcttaa gtctagaggg cccgtttaaa cccgctgatc agcctcgact
     2521 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     2581 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     2641 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     2701 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggctcgagg gggttggggt
     2761 tgcgcctttt ccaaggcagc cctgggtttg cgcagggacg cggctgctct gggcgtggtt
     2821 ccgggaaacg cagcggcgcc gaccctggga ctcgcacatt cttcacgtcc gttcgcagcg
     2881 tcacccggat cttcgccgct acccttgtgg gccccccggc gacgcttcct gctccgcccc
     2941 taagtcggga aggttccttg cggttcgcgg cgtgccggac gtgacaaacg gaagccgcac
     3001 gtctcactag taccctcgca gacggacagc gccagggagc aatggcagcg cgccgaccgc
     3061 gatgggctgt ggccaatagc ggctgctcag cagggcgcgc cgagagcagc ggccgggaag
     3121 gggcggtgcg ggaggcgggg tgtggggcgg tagtgtgggc cctgttcctg cccgcgcggt
     3181 gttccgcatt ctgcaagcct ccggagcgca cgtcggcagt cggctccctc gttgaccgaa
     3241 tcaccgacct ctctccccag ggggatccac cggagcttac catgaccgag tacaagccca
     3301 cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt
     3361 tcgccgacta ccccgccacg cgccacaccg tcgatccgga ccgccacatc gagcgggtca
     3421 ccgagctgca agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg
     3481 cggacgacgg cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg
     3541 tgttcgccga gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc
     3601 aacagatgga aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca
     3661 ccgtcggcgt ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg
     3721 gagtggaggc ggccgagcgc gccggggtgc ccgccttcct ggaaacctcc gcgccccgca
     3781 acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag
     3841 gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg acgcccgccc cacgacccgc
     3901 agcgcccgac cgaaaggagc gcacgacccc atgcatcggt acctttaaga ccaatgactt
     3961 acaaggcagc tgtagatctt agccactttc tagagtcggg gcggccggcc gcttcgagca
     4021 gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa
     4081 tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat
     4141 aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg
     4201 gaggtttttt aaagcaagta aaacctctac aaatgtggta aaatcgataa ggatccgtcg
     4261 accgatgccc ttgagagcct tcaacccagt cagctccttc cggtgggcgc ggggcatgac
     4321 tatcgtcgcc gcacttatga ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc
     4381 agcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttagg ctgcggcgag
     4441 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     4501 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     4561 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     4621 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     4681 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     4741 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     4801 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     4861 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     4921 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     4981 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     5041 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     5101 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     5161 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     5221 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     5281 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     5341 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     5401 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     5461 taccgcggga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     5521 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     5581 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     5641 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     5701 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     5761 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     5821 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     5881 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     5941 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     6001 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     6061 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     6121 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     6181 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     6241 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     6301 cccgaaaagt gccacctg
//
