LOCUS       sequence-405617-        6261 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A backbone containing a RsDddA fragment B for constructing 
            mitochondria-targeted RsDdCBE expression plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6261)
  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 6261)
  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-B-UGI-PURO
FEATURES             Location/Qualifiers
     source          1..6261
                     /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             2169..2423
                     /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(2456..2473)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2462..2686
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        2693..3203
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3122..3140
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3225..3824
                     /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(3225..3244)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3721..3741
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3971..4092
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3982..4001)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4036..4055
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4340..4357)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4511..5099)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4591..4610)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5270..6061,6062..6130))
                     /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     5893..5912
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6131..6235)
                     /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 ccgctagcag cgtggccaac aacgtgagag
     2101 ccatccccgt gcccaagacc tacatcggca acagcaccgt gcccaagatc aaggcatcag
     2161 gcgggagcac caacctgtct gacatcatcg agaaggagac aggcaagcag ctggtcatcc
     2221 aggagagcat cctgatgctg cccgaagaag tcgaagaagt gatcggaaac aagcctgaga
     2281 gcgatatcct ggtccatacc gcctacgacg agagtaccga cgaaaatgtg atgctgctga
     2341 catccgacgc cccagagtat aagccctggg ctctggtcat ccaggattcc aacggagaga
     2401 acaaaatcaa aatgctggct taagtctaga gggcccgttt aaacccgctg atcagcctcg
     2461 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     2521 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
     2581 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
     2641 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggctcg agggggttgg
     2701 ggttgcgcct tttccaaggc agccctgggt ttgcgcaggg acgcggctgc tctgggcgtg
     2761 gttccgggaa acgcagcggc gccgaccctg ggactcgcac attcttcacg tccgttcgca
     2821 gcgtcacccg gatcttcgcc gctacccttg tgggcccccc ggcgacgctt cctgctccgc
     2881 ccctaagtcg ggaaggttcc ttgcggttcg cggcgtgccg gacgtgacaa acggaagccg
     2941 cacgtctcac tagtaccctc gcagacggac agcgccaggg agcaatggca gcgcgccgac
     3001 cgcgatgggc tgtggccaat agcggctgct cagcagggcg cgccgagagc agcggccggg
     3061 aaggggcggt gcgggaggcg gggtgtgggg cggtagtgtg ggccctgttc ctgcccgcgc
     3121 ggtgttccgc attctgcaag cctccggagc gcacgtcggc agtcggctcc ctcgttgacc
     3181 gaatcaccga cctctctccc cagggggatc caccggagct taccatgacc gagtacaagc
     3241 ccacggtgcg cctcgccacc cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg
     3301 cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg
     3361 tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg
     3421 tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg
     3481 cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc
     3541 agcaacagat ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg tggttcctgg
     3601 ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc
     3661 ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt cctggaaacc tccgcgcccc
     3721 gcaacctccc cttctacgag cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg
     3781 aaggaccgcg cacctggtgc atgacccgca agcccggtgc ctgacgcccg ccccacgacc
     3841 cgcagcgccc gaccgaaagg agcgcacgac cccatgcatc ggtaccttta agaccaatga
     3901 cttacaaggc agctgtagat cttagccact ttctagagtc ggggcggccg gccgcttcga
     3961 gcagacatga taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa
     4021 aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc
     4081 aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggaggtg
     4141 tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg gtaaaatcga taaggatccg
     4201 tcgaccgatg cccttgagag ccttcaaccc agtcagctcc ttccggtggg cgcggggcat
     4261 gactatcgtc gccgcactta tgactgtctt ctttatcatg caactcgtag gacaggtgcc
     4321 ggcagcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt aggctgcggc
     4381 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     4441 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     4501 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     4561 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     4621 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     4681 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     4741 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     4801 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     4861 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     4921 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     4981 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     5041 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     5101 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     5161 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     5221 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     5281 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     5341 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     5401 tgataccgcg ggacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     5461 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     5521 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     5581 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     5641 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     5701 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     5761 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     5821 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     5881 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     5941 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     6001 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     6061 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     6121 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     6181 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     6241 ttccccgaaa agtgccacct g
//
