LOCUS       sequence-225037-        2457 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  POC12371, MetClo assembly vector with p15A replication origin and 
            kanamycin resistance for BsaI-based MetClo, adaptor sequence type 
            d-e.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2457)
  AUTHORS   Lin D, O'Callaghan CA
  TITLE     MetClo: methylase-assisted hierarchical DNA assembly using a single 
            type IIS restriction enzyme.
  JOURNAL   Nucleic Acids Res. 2018 Jul 9. pii: 5050631. doi: 
            10.1093/nar/gky596.
  PUBMED    29986052
REFERENCE   2  (bases 1 to 2457)
  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: "Nucleic 
            Acids Res. 2018 Jul 9. pii: 5050631. doi: 10.1093/nar/gky596."
COMMENT     Sequence Label: pMXLK_dFe
FEATURES             Location/Qualifiers
     source          1..2457
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    111..132
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(147..152,153..170,171..177)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    185..201
                     /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)."
     primer_bind     190..212
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     209..225
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     209..225
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             221..544
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    238..294
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(295..312)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(295..311)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(304..326)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(520..539)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        707..811
                     /gene="bla"
                     /label=AmpR promoter
     CDS             812..1606
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSLREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     rep_origin      1632..2177
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     2078..2097
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2294..2311
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cactctgtga cttggtctca acgaggagtg agaccggcct agcaggtgtg agtttgcagc
       61 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt
      121 tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt
      181 ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag
      241 cttgcatgcc tgcaggtcga ctctagagga tccccgggta ccgagctcga attcactggc
      301 cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc
      361 agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc
      421 ccaacagttg cgcagcctga atggcgaatg gcgcctgatg cggtattttc tccttacgca
      481 tctgtgcggt atttcacacc gcatatggtg cactctcagt acaatctgct ctgatgccgc
      541 atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
      601 gctcccggca tccgcttaca gacaagctgt gacaagtacc acctggaacg ccaaaggtct
      661 cacgctcaac tgagaccaag tcacgaagtg tgcggccgcc ttcgtgcgcg gaacccctat
      721 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
      781 aatgcttcaa taatattgaa aaaggaagag tatggctaaa atgagaatat caccggaatt
      841 gaaaaaactg atcgaaaaat accgctgcgt aaaagatacg gaaggaatgt ctcctgctaa
      901 ggtatataag ctggtgggag aaaatgaaaa cctatattta aaaatgacgg acagccggta
      961 taaagggacc acctatgatg tggaacggga aaaggacatg atgctatggc tggaaggaaa
     1021 gctgcctgtt ccaaaggtcc tgcactttga acggcatgat ggctggagca atctgctcat
     1081 gagtgaggcc gatggcgtcc tttgctcgga agagtatgaa gatgaacaaa gccctgaaaa
     1141 gattatcgag ctgtatgcgg agtgcatcag gctctttcac tccatcgaca tatcggattg
     1201 tccctatacg aatagcttag acagccgctt agccgaattg gattacttac tgaataacga
     1261 tctggccgat gtggattgcg aaaactggga agaggacact ccatttaaag atccgcgcga
     1321 gctgtatgat tttttaaaga cggaaaagcc cgaagaggaa cttgtctttt cccacggcga
     1381 cctgggagac agcaacatct ttgtgaaaga tggcaaagta agtggcttta ttgatcttgg
     1441 gagaagcggc agggcggaca agtggtatga cattgccttc tgcgtccggt cgctcaggga
     1501 ggatatcggg gaagaacagt atgtcgagct attttttgac ttactgggga tcaagcctga
     1561 ttgggagaaa ataaaatatt atattttact ggatgaattg ttttaggagc ctgtttaata
     1621 agatgatctt cttgagatcg ttttggtctg cgcgtaatct cttgctctga aaacgaaaaa
     1681 accgccttgc agggcggttt ttcgaaggtt ctctgagcta ccaactcttt gaaccgaggt
     1741 aactggcttg gaggagcgca gtcaccaaaa cttgtccttt cagtttagcc ttaaccggcg
     1801 catgacttca agactaactc ctctaaatca attaccagtg gctgctgcca gtggtgcttt
     1861 tgcatgtctt tccgggttgg actcaagacg atagttaccg gataaggcgc agcggtcgga
     1921 ctgaacgggg ggttcgtgca tacagtccag cttggagcga actgcctacc cggaactgag
     1981 tgtcaggcgt ggaatgagac aaacgcggcc ataacagcgg aatgacaccg gtaaaccgaa
     2041 aggcaggaac aggagagcgc acgagggagc cgccaggggg aaacgcctgg tatctttata
     2101 gtcctgtcgg gtttcgccac cactgatttg agcgtcagat ttcgtgatgc ttgtcagggg
     2161 ggcggagcct atggaaaaac ggctttgccg cggccctctc acttccctgt taagtatctt
     2221 cctggcatct tccaggaaat ctccgccccg ttcgtaagcc atttccgctc gccgcagtcg
     2281 aacgaccgag cgtagcgagt cagtgagcga ggaagcggaa tatatcctgt atcacatatt
     2341 ctgctgacgc accggtgcag ccttttttct cctgccacat gaagcacttc actgacaccc
     2401 tcatcagtgc caacatagta agccagtata cactccgcta gcgcgcaagc ggccgca
//
