LOCUS       sequence-457184-        4092 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for MLH1-SB overexpression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4092)
  AUTHORS   Park JC, Uhm H, Kim YW, Oh YE, Lee JH, Yang J, Kim K, Bae S
  TITLE     AI-generated MLH1 small binder improves prime editing efficiency.
  JOURNAL   Cell. 2025 Aug 1:S0092-8674(25)00799-8. doi: 
            10.1016/j.cell.2025.07.010.
  PUBMED    40769155
REFERENCE   2  (bases 1 to 4092)
  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: "Cell. 2025 
            Aug 1:S0092-8674(25)00799-8. doi: 10.1016/j.cell.2025.07.010."
COMMENT     Sequence Label: pCMV-PuroR-2A-MLH1-SB
FEATURES             Location/Qualifiers
     source          1..4092
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     protein_bind    158..205
                     /label=Bxb1 attP
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attP site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     enhancer        273..652
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        653..856
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     807..827
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     853..877
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        973..991
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1005..1601
                     /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(1005..1024)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1501..1521
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             1602..1658
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     promoter        complement(1946..1964)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1947..1964)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1984..2001)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1990..2214
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2302..2319)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2473..3061)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2553..2572)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3232..4023,4024..4092))
                     /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     3855..3874
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct gacgtcgaga attcgtttaa acctcgaggt ttgtctggtc aaccaccgcg
      181 gtctcagtgg tgtacggtac aaaccgcggc cgcgttaagc cagtatctgc tccctgcttg
      241 tgtgttggag aaccagagcc acattaaccg ctgacattga ttattgacta gttattaata
      301 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
      361 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
      421 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
      481 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc
      541 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg
      601 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg
      661 gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
      721 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
      781 atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt
      841 ctatataagc agagctcgtt tagtgaaccg tcagatcgcc tggaggcgcc atccacgctg
      901 ttttgacctc catagaagac accgggaccg atccagcctc cggactctag cctaggcttt
      961 tgcaaaaagc tctaatacga ctcactatag gcaagcttgc caccatgacc gagtacaagc
     1021 ccacggtgcg cctcgccacc cgcgacgacg tcccccgggc cgtacgcacc ctcgccgccg
     1081 cgttcgccga ctaccccgcc acgcgccaca ccgtcgaccc ggaccgccac atcgagcggg
     1141 tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg
     1201 tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg
     1261 cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc
     1321 agcaacagat ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg tggttcctgg
     1381 ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc
     1441 ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc
     1501 gcaacctccc cttctacgag cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg
     1561 aaggaccgcg cacctggtgc atgacccgca agcccggtgc cgctactaac ttcagcctgc
     1621 tgaagcaggc tggagacgtg gaggagaacc ctggacctgg taccatgagc gaggaagagg
     1681 agaaaagaaa gatcgaggaa ggcatgcgga gactgagaga gaacctgaaa aagaagcgga
     1741 aggaagagga agagaaggag aagaaggaaa ccgaggagct ggctaaactg gccaaggaaa
     1801 tcgtggaaga ggccaagaaa gagctgaagg ccgagggcaa gtggaagaac aaggaagaga
     1861 tctgggaggc cttcaagaag aaaatcctgg aaaagttcga gaagctggaa taaaagcttt
     1921 cgagcatgca tctagagggc cctattctat agtgtcacct aaatgctaga gctcgctgat
     1981 cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt
     2041 ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat
     2101 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg
     2161 gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg
     2221 aggcggaaag aaccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     2281 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     2341 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     2401 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     2461 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     2521 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     2581 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     2641 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     2701 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     2761 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     2821 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     2881 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     2941 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     3001 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     3061 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     3121 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     3181 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     3241 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     3301 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     3361 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     3421 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     3481 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     3541 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     3601 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     3661 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     3721 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     3781 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     3841 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     3901 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     3961 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     4021 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     4081 ttgaatactc at
//
