LOCUS       sequence-343632-        6315 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of mouse MLH1dn (codon optimized).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6315)
  AUTHORS   Chen PJ, Hussmann JA, Yan J, Knipping F, Ravisankar P, Chen PF, Chen
            C, Nelson JW, Newby GA, Sahin M, Osborn MJ, Weissman JS, Adamson B, 
            Liu DR
  TITLE     Enhanced prime editing systems by manipulating cellular determinants
            of editing outcomes.
  JOURNAL   Cell. 2021 Oct 28;184(22):5635-5652.e29. doi: 
            10.1016/j.cell.2021.09.018.
  PUBMED    34653350
REFERENCE   2  (bases 1 to 6315)
  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"; doi: 
            "10.1016/j.cell.2021.09"; journalName: "Cell"; date: "2021-10-28- 
            28"; volume: "184"; issue: "22"; pages: "5635-5652.e29"
COMMENT     Sequence Label: pEF1a-mMLH1dn
FEATURES             Location/Qualifiers
     source          1..6315
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        197..1375
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          428..1366
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1323..1343
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1394..1403
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3679..3696
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3719..3736)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3725..3949
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4020..4036)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4020..4036)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4033..4055)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4044..4060
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4068..4074,4075..4092,4093..4098))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4113..4134
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4251..4268)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4422..5010)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4502..4521)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5181..5972,5973..6041))
                     /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     5804..5823
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6042..6146)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6225..6244)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt
       61 tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt gacattgatt attgactgca
      121 aagatggata aagttttaaa cagagaggaa tctttgcagc taatggacct tctaggtctt
      181 gaaaggagtg ggaattggct ccggtgcccg tcagtgggca gagcgcacat cgcccacagt
      241 ccccgagaag ttggggggag gggtcggcaa ttgaaccggt gcctagagaa ggtggcgcgg
      301 ggtaaactgg gaaagtgatg tcgtgtactg gctccgcctt tttcccgagg gtgggggaga
      361 accgtatata agtgcagtag tcgccgtgaa cgttcttttt cgcaacgggt ttgccgccag
      421 aacacaggta agtgccgtgt gtggttcccg cgggcctggc ctctttacgg gttatggccc
      481 ttgcgtgcct tgaattactt ccacctggct gcagtacgtg attcttgatc ccgagcttcg
      541 ggttggaagt gggtgggaga gttcgaggcc ttgcgcttaa ggagcccctt cgcctcgtgc
      601 ttgagttgag gcctggcctg ggcgctgggg ccgccgcgtg cgaatctggt ggcaccttcg
      661 cgcctgtctc gctgctttcg ataagtctct agccatttaa aatttttgat gacctgctgc
      721 gacgcttttt ttctggcaag atagtcttgt aaatgcgggc caagatctgc acactggtat
      781 ttcggttttt ggggccgcgg gcggcgacgg ggcccgtgcg tcccagcgca catgttcggc
      841 gaggcggggc ctgcgagcgc ggccaccgag aatcggacgg gggtagtctc aagctggccg
      901 gcctgctctg gtgcctggcc tcgcgccgcc gtgtatcgcc ccgccctggg cggcaaggct
      961 ggcccggtcg gcaccagttg cgtgagcgga aagatggccg cttcccggcc ctgctgcagg
     1021 gagctcaaaa tggaggacgc ggcgctcggg agagcgggcg ggtgagtcac ccacacaaag
     1081 gaaaagggcc tttccgtcct cagccgtcgc ttcatgtgac tccacggagt accgggcgcc
     1141 gtccaggcac ctcgattagt tctcgagctt ttggagtacg tcgtctttag gttgggggga
     1201 ggggttttat gcgatggagt ttccccacac tgagtgggtg gagactgaag ttaggccagc
     1261 ttggcacttg atgtaattct ccttggaatt tgcccttttt gagtttggat cttggttcat
     1321 tctcaagcct cagacagtgg ttcaaagttt ttttcttcca tttcaggtgt cgtgacgtac
     1381 gggatcctct agagccacca tggcattcgt ggctggtgtc atccgccgtc ttgatgagac
     1441 tgtcgtcaac agaattgccg caggagaagt catccagcgt ccagccaacg ccatcaagga
     1501 aatgatcgaa aactgtctgg atgccaagag cacaaacata caagttgtgg tgaaggaggg
     1561 tggcctcaaa ctcattcaga tccaagacaa cggcacggga attagaaaag aagacctgga
     1621 cattgtgtgc gagcgcttca caacctctaa attgcagaca tttgaggacc tggcaagcat
     1681 cagtacttat ggcttccgag gggaggctct ggcctccata agccatgtgg ctcacgtgac
     1741 catcacgaca aaaacggctg atggaaagtg cgcataccgt gcatcctact cggatgggaa
     1801 actacaggcc cctccaaaac cttgtgccgg caaccagggc acactcatca ctgttgaaga
     1861 tttgttctac aacattatca cacggcggaa ggctttaaag aatccaagtg aggaatatgg
     1921 gaagatcctt gaagtggtcg gacgatactc tatccacaac tctgggattt ccttctctgt
     1981 gaagaagcaa ggtgaaactg tttctgatgt gcgcaccttg cccaatgcta ccactgtgga
     2041 caatattcgc agcatctttg ggaatgcggt ctctcgagag ctcattgagg ttggatgtga
     2101 ggacaagaca ctggctttca agatgaatgg ctacatttcc aatgccaact acagcgttaa
     2161 aaagtgtatc ttcctgctgt ttatcaacca taggctggtg gagtcagctg cactccgaaa
     2221 agccatagag acagtatatg ctgcatactt gcccaaaaac acgcatccat tcctctacct
     2281 gtcgctagaa atctcccctc agaacgtcga tgtcaatgtc caccccacca agcatgaagt
     2341 tcacttcctt catgaggaat caatactgca aagagtgcag caacatatag aaagcaaatt
     2401 actgggttcc aacagttcaa gaatgtattt cacccagact cttctgcctg gtctggcagg
     2461 ccctagtgga gaagcagcac ggccgaccac tggagtggcc tcaagcagta ccagtggttc
     2521 tggagacaaa gtgtatgcct accagatggt caggacagac tctcgggagc agaagctcga
     2581 cgctttttta cagcctgtgt caagcctggg accctcccag ccacaagatc ctgccccggt
     2641 aaggggagcc aggacagaag gctccccaga gcgggccacg agagaggatg aagagatgct
     2701 ggcgctccca gcgccagctg aggccgcagc tgaatctgag aatctggaga gagagtctct
     2761 gatggagacc tccgatgcag cccagaaagc tgcacccaca tcatctccag gctcttcaag
     2821 gaaaaggcac agagaagact ctgatgtaga gatggtcgaa aatgcatcag ggaaagaaat
     2881 gacagctgcc tgctacccgc gtaggagaat tataaacctg acaagtgtgt tatcgttgca
     2941 ggaggagatt agtgaaaggt gccacgaaac cttgagagag atgctgagga atcatagctt
     3001 tgtgggctgt gtgaaccccc aatgggccct ggctcagcac cagaccaaac tatatttgct
     3061 taatactacc aagctgtcag aagaactgtt ctatcagata ctcatttatg actttgccaa
     3121 ctttggagtt ttgcggctga gcgagcccgc gccactcttt gacctagcca tgcttgctct
     3181 ggacagccct gaaagtggct ggactgaaga tgacggcccg aaggaagggt tggctgagta
     3241 tattgtagaa tttctgaaga agaaagcaga gatgctcgcc gactacttct ccgtagagat
     3301 cgacgaggag ggcaatctta tcggattacc tctcctcata gactcttatg tgcccccttt
     3361 ggaggggctg cccattttca tcctccgcct cgctactgaa gtaaactggg atgaggagaa
     3421 ggagtgcttc gagagtctta gtaaagagtg tgcaatgttt tatagcatta ggaagcagta
     3481 catcctggaa gagtccacac tgtctgggca gcagtcagac atgcctggta gcaccagcaa
     3541 gccatggaag tggactgtgg agcacatcat ctacaaggcc ttccggtccc acctgcttcc
     3601 tcccaagcac tttaccgagg atgggaatgt gctccagctg gccaatctcc ctgatctgta
     3661 caaagttttt taaccggtca tcatcaccat caccattgag tttaaacccg ctgatcagcc
     3721 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     3781 accctggaag gtgccactcc cactgtcctt tcctaataaa atgagaaaat tgcatcgcat
     3841 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
     3901 gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg
     3961 gaaagaacca gctggggctc gataccgtcg acctctagct agagcttggc gtaatcatgg
     4021 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     4081 ggaagcataa agtgtaaagc ctagggtgcc taatgagtga gctaactcac attaattgcg
     4141 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     4201 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     4261 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     4321 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     4381 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     4441 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     4501 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     4561 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     4621 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     4681 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     4741 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     4801 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     4861 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     4921 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     4981 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     5041 gacactcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     5101 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     5161 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     5221 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     5281 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     5341 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     5401 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     5461 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     5521 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     5581 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     5641 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     5701 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     5761 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     5821 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     5881 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     5941 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     6001 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     6061 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     6121 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac
     6181 ggatcgggag atcgatctcc cgatccccta gggtcgactc tcagtacaat ctgctctgat
     6241 gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc
     6301 gcgagcaaaa tttaa
//
