LOCUS       sequence-350215-        6303 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human MLH1dn (original codon usage).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6303)
  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 6303)
  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-hMLH1dn (og codon)
FEATURES             Location/Qualifiers
     source          1..6303
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        192..1370
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          423..1361
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1318..1338
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             3662..3679
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3702..3719)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3708..3932
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4003..4019)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4003..4019)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4016..4038)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4027..4043
                     /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(4051..4057,4058..4075,4076..4081))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4096..4117
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4234..4251)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4405..4993)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4485..4504)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5164..5955,5956..6024))
                     /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     5787..5806
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6025..6129)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6208..6227)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aacaaggcaa ggcttgaccg acaattgcat gaagaatctg cttagggtta ggcgttttgc
       61 gctgcttcgc gatgtacggg ccagatatac gcgttgacat tgattattga ctgcaaagat
      121 ggataaagtt ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag
      181 gagtgggaat tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg
      241 agaagttggg gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa
      301 actgggaaag tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt
      361 atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac
      421 aggtaagtgc cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg
      481 tgccttgaat tacttccacc tggctgcagt acgtgattct tgatcccgag cttcgggttg
      541 gaagtgggtg ggagagttcg aggccttgcg cttaaggagc cccttcgcct cgtgcttgag
      601 ttgaggcctg gcctgggcgc tggggccgcc gcgtgcgaat ctggtggcac cttcgcgcct
      661 gtctcgctgc tttcgataag tctctagcca tttaaaattt ttgatgacct gctgcgacgc
      721 tttttttctg gcaagatagt cttgtaaatg cgggccaaga tctgcacact ggtatttcgg
      781 tttttggggc cgcgggcggc gacggggccc gtgcgtccca gcgcacatgt tcggcgaggc
      841 ggggcctgcg agcgcggcca ccgagaatcg gacgggggta gtctcaagct ggccggcctg
      901 ctctggtgcc tggcctcgcg ccgccgtgta tcgccccgcc ctgggcggca aggctggccc
      961 ggtcggcacc agttgcgtga gcggaaagat ggccgcttcc cggccctgct gcagggagct
     1021 caaaatggag gacgcggcgc tcgggagagc gggcgggtga gtcacccaca caaaggaaaa
     1081 gggcctttcc gtcctcagcc gtcgcttcat gtgactccac ggagtaccgg gcgccgtcca
     1141 ggcacctcga ttagttctcg agcttttgga gtacgtcgtc tttaggttgg ggggaggggt
     1201 tttatgcgat ggagtttccc cacactgagt gggtggagac tgaagttagg ccagcttggc
     1261 acttgatgta attctccttg gaatttgccc tttttgagtt tggatcttgg ttcattctca
     1321 agcctcagac agtggttcaa agtttttttc ttccatttca ggtgtcgtga cgtacgggat
     1381 cctctagagc caccatgtcg ttcgtggcag gggttattcg gcggctggac gagacagtgg
     1441 tgaaccgcat cgcggcgggg gaagttatcc agcggccagc taatgctatc aaagagatga
     1501 ttgagaactg tttagatgca aaatccacaa gtattcaagt gattgttaaa gagggaggcc
     1561 tgaagttgat tcagatccaa gacaatggca ccgggatcag gaaagaagat ctggatattg
     1621 tatgtgaaag gttcactact agtaaactgc agtcctttga ggatttagcc agtatttcta
     1681 cctatggctt tcgaggtgag gctttggcca gcataagcca tgtggctcat gttactatta
     1741 caacgaaaac agctgatgga aagtgtgcat acagagcaag ttactcagat ggaaaactga
     1801 aagcccctcc taaaccatgt gctggcaatc aagggaccca gatcacggtg gaggaccttt
     1861 tttacaacat agccacgagg agaaaagctt taaaaaatcc aagtgaagaa tatgggaaaa
     1921 ttttggaagt tgttggcagg tattcagtac acaatgcagg cattagtttc tcagttaaaa
     1981 aacaaggaga gacagtagct gatgttagga cactacccaa tgcctcaacc gtggacaata
     2041 ttcgctccat ctttggaaat gctgttagtc gagaactgat agaaattgga tgtgaggata
     2101 aaaccctagc cttcaaaatg aatggttaca tatccaatgc aaactactca gtgaagaagt
     2161 gcatcttctt actcttcatc aaccatcgtc tggtagaatc aacttccttg agaaaagcca
     2221 tagaaacagt gtatgcagcc tatttgccca aaaacacaca cccattcctg tacctcagtt
     2281 tagaaatcag tccccagaat gtggatgtta atgtgcaccc cacaaagcat gaagttcact
     2341 tcctgcacga ggagagcatc ctggagcggg tgcagcagca catcgagagc aagctcctgg
     2401 gctccaattc ctccaggatg tacttcaccc agactttgct accaggactt gctggcccct
     2461 ctggggagat ggttaaatcc acaacaagtc tgacctcgtc ttctacttct ggaagtagtg
     2521 ataaggtcta tgcccaccag atggttcgta cagattcccg ggaacagaag cttgatgcat
     2581 ttctgcagcc tctgagcaaa cccctgtcca gtcagcccca ggccattgtc acagaggata
     2641 agacagatat ttctagtggc agggctaggc agcaagatga ggagatgctt gaactcccag
     2701 cccctgctga agtggctgcc aaaaatcaga gcttggaggg ggatacaaca aaggggactt
     2761 cagaaatgtc agagaagaga ggacctactt ccagcaaccc cagaaagaga catcgggaag
     2821 attctgatgt ggaaatggtg gaagatgatt cccgaaagga aatgactgca gcttgtaccc
     2881 cccggagaag gatcattaac ctcactagtg ttttgagtct ccaggaagaa attaatgagc
     2941 agggacatga ggttctccgg gagatgttgc ataaccactc cttcgtgggc tgtgtgaatc
     3001 ctcagtgggc cttggcacag catcaaacca agttatacct tctcaacacc accaagctta
     3061 gtgaagaact gttctaccag atactcattt atgattttgc caattttggt gttctcaggt
     3121 tatcggagcc agcaccgctc tttgaccttg ccatgcttgc cttagatagt ccagagagtg
     3181 gctggacaga ggaagatggt cccaaagaag gacttgctga atacattgtt gagtttctga
     3241 agaagaaggc tgagatgctt gcagactatt tctctttgga aattgatgag gaagggaacc
     3301 tgattggatt accccttctg attgacaact atgtgccccc tttggaggga ctgcctatct
     3361 tcattcttcg actagccact gaggtgaatt gggacgaaga aaaggaatgt tttgaaagcc
     3421 tcagtaaaga atgcgctatg ttctattcca tccggaagca gtacatatct gaggagtcga
     3481 ccctctcagg ccagcagagt gaagtgcctg gctccattcc aaactcctgg aagtggactg
     3541 tggaacacat tgtctataaa gccttgcgct cacacattct gcctcctaaa catttcacag
     3601 aagatggaaa tatcctgcag cttgctaacc tgcctgatct atacaaagtc ttttaaccgg
     3661 tcatcatcac catcaccatt gagtttaaac ccgctgatca gcctcgactg tgccttctag
     3721 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     3781 tcccactgtc ctttcctaat aaaatgagaa aattgcatcg cattgtctga gtaggtgtca
     3841 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     3901 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     3961 ctcgataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     4021 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     4081 agcctagggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     4141 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4201 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4261 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4321 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4381 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4441 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4501 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4561 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4621 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4681 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4741 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     4801 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     4861 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     4921 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     4981 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacactc agtggaacga
     5041 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     5101 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5161 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5221 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5281 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5341 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5401 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     5461 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5521 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5581 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5641 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     5701 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     5761 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     5821 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     5881 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     5941 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6001 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     6061 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6121 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatcgatc
     6181 tcccgatccc ctagggtcga ctctcagtac aatctgctct gatgccgcat agttaagcca
     6241 gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc
     6301 tac
//
