LOCUS       sequence-225138-        7974 bp    DNA     circular SYN 15-APR-2025
DEFINITION  POC12394, MetClo assembly vector with F replication origin and 
            chloramphenicol resistance for BsaI-based MetClo, adaptor sequence 
            type a-q.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7974)
  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 7974)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: pMXBC_aFq
FEATURES             Location/Qualifiers
     source          1..7974
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      224..812
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     713..732
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     966..983
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1100..1121
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1136..1141,1142..1159,1160..1166)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1174..1190
                     /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     1179..1201
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1198..1214
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1198..1214
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1210..1533
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    1227..1283
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(1284..1301)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1284..1300)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1293..1315)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1509..1528)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        1804..1906
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             1907..2566
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1973..1992)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     rep_origin      3381..3600
                     /label=ori2
                     /note="secondary origin of replication for the bacterial F 
                     plasmid; also known as oriS"
     CDS             3691..4446
                     /codon_start=1
                     /gene="repE"
                     /product="replication initiation protein for the bacterial 
                     F plasmid"
                     /label=repE
                     /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ
                     IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG
                     DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM
                     RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM
                     RLSYIEKKKGRQTTHIVFSFRDITSMTTG"
     misc_feature    4449..4699
                     /gene="incC"
                     /label=incC
                     /note="incompatibility region of the bacterial F plasmid"
     CDS             5025..6200
                     /codon_start=1
                     /gene="sopA"
                     /product="partitioning protein for the bacterial F plasmid"
                     /label=sopA
                     /translation="MFRMKLMETLNQCINAGHEMTKAIAIAQFNDDSPEARKITRRWRI
                     GEAADLVGVSSQAIRDAEKAGRLPHPDMEIRGRVEQRVGYTIEQINHMRDVFGTRLRRA
                     EDVFPPVIGVAAHKGGVYKTSVSVHLAQDLALKGLRVLLVEGNDPQGTASMYHGWVPDL
                     HIHAEDTLLPFYLGEKDDVTYAIKPTCWPGLDIIPSCLALHRIETELMGKFDEGKLPTD
                     PHLMLRLAIETVAHDYDVIVIDSAPNLGIGTINVVCAADVLIVPTPAELFDYTSALQFF
                     DMLRDLLKNVDLKGFEPDVRILLTKYSNSNGSQSPWMEEQIRDAWGSMVLKNVVRETDE
                     VGKGQIRMRTVFEQAIDQRSSTGAWRNALSIWEPVCNEIFDRLIKPRWEIR"
     CDS             6200..7171
                     /codon_start=1
                     /gene="sopB"
                     /product="partitioning protein for the bacterial F plasmid"
                     /label=sopB
                     /translation="MKRAPVIPKHTLNTQPVEDTSLSTPAAPMVDSLIARVGVMARGNA
                     ITLPVCGRDVKFTLEVLRGDSVEKTSRVWSGNERDQELLTEDALDDLIPSFLLTGQQTP
                     AFGRRVSGVIEIADGSRRRKAAALTESDYRVLVGELDDEQMAALSRLGNDYRPTSAYER
                     GQRYASRLQNEFAGNISALADAENISRKIITRCINTAKLPKSVVALFSHPGELSARSGD
                     ALQKAFTDKEELLKQQASNLHEQKKAGVIFEAEEVITLLTSVLKTSSASRTSLSSRHQF
                     APGATVLYKGDKMVLNLDRSRVPTECIEKIEAILKELEKPAP"
     misc_feature    7244..7717
                     /gene="sopC"
                     /label=sopC
                     /note="centromere-like partitioning region of the bacterial
                     F plasmid"
ORIGIN
        1 cactctgtga cttggtctca aggtggagtg agaccggcct agcaggtgtg agtttgtcag
       61 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
      121 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
      181 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
      241 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
      301 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
      361 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
      421 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
      481 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
      541 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
      601 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
      661 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
      721 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
      781 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
      841 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
      901 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
      961 agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc
     1021 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg
     1081 gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc caggctttac
     1141 actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag
     1201 gaaacagcta tgaccatgat tacgccaagc ttgcatgcct gcaggtcgac tctagaggat
     1261 ccccgggtac cgagctcgaa ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac
     1321 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat
     1381 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     1441 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg catatggtgc
     1501 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca
     1561 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     1621 acaagtacca cctggaacgc caaaggtctc acgcttgaga ccaagtcacg aagtgtgcgg
     1681 ccgccttcag cggccgcacg aataaatacc tgtgacggaa gatcacttcg cagaataaat
     1741 aaatcctggt gtccctgttg ataccgggaa gccctgggcc aacttttggc gaaaatgaga
     1801 cgttgatcgg cacgtaagag gttccaactt tcaccataat gaaataagat cactaccggg
     1861 cgtatttttt gagttatcga gattttcagg agctaaggaa gctaaaatgg agaaaaaaat
     1921 cactggatat accaccgttg atatatccca atggcatcgt aaagaacatt ttgaggcatt
     1981 tcagtcagtt gctcaatgta cctataacca gaccgttcag ctggatatta cggccttttt
     2041 aaagaccgta aagaaaaata agcacaagtt ttatccggcc tttattcaca ttcttgcccg
     2101 cctgatgaat gctcatccgg agttccgtat ggcaatgaaa gacggtgagc tggtgatatg
     2161 ggatagtgtt cacccttgtt acaccgtttt ccatgagcaa actgaaacgt tttcatcgct
     2221 ctggagtgaa taccacgacg atttccggca gtttctacac atatattcgc aagatgtggc
     2281 gtgttacggt gaaaacctgg cctatttccc taaagggttt attgagaata tgtttttcgt
     2341 ctcagccaat ccctgggtga gtttcaccag ttttgattta aacgtggcca atatggacaa
     2401 cttcttcgcc cccgttttca ccatgggcaa atattatacg caaggcgaca aggtgctgat
     2461 gccgctggcg attcaggttc atcatgccgt ttgtgatggc ttccatgtcg gcagaatgct
     2521 taatgaatta caacagtact gcgatgagtg gcagggcggg gcgtaagagc tggagcggcg
     2581 taaccgtcgc acaggaagga cagagaaagc gcggatctgg gaagtgacgg acagaacggt
     2641 caggacctgg attggggagg cggttgccgc cgctgctgct gacggtgtga cgttctctgt
     2701 tccggtcaca ccacatacgt tccgccattc ctatgcgatg cacatgctgt atgccggtat
     2761 accgctgaaa gttctgcaaa gcctgatggg acataagtcc atcagttcaa cggaagtcta
     2821 cacgaaggtt tttgcgctgg atgtggctgc ccggctccgg gtgcagtttg cgatgccgga
     2881 gtctgatgcg gttgcgatgc tgaaacaatt atcctgagaa taaatgcctt ggcctttata
     2941 tggaaatgtg gaactgagtg gatatgctgt ttttgtctgt taaacagaga agctggctgt
     3001 tatccactga gaagcgaacg aaacagtcgg gaaaatctcc cattatcgta gagatccgca
     3061 ttattaatct caggagcctg tgtagcgttt ataggaagta gtgttctgtc atgatgcctg
     3121 caagcggtaa cgaaaacgat ttgaatatgc cttcaggaac aatagaaatc ttcgtgcggt
     3181 gttacgttga agtggagcgg attatgtcag caatggacag aacaacctaa tgaacacaga
     3241 accatgatgt ggtctgtcct tttacagcca gtagtgctcg ccgcagtcga gcgacagggc
     3301 gaagccctcg agtgagcgag gaagcaccag ggaacagcac ttatatattc tgcttacaca
     3361 cgatgcctga aaaaacttcc cttggggtta tccacttatc cacggggata tttttataat
     3421 tatttttttt atagttttta gatcttcttt tttagagcgc cttgtaggcc tttatccatg
     3481 ctggttctag agaaggtgtt gtgacaaatt gccctttcag tgtgacaaat caccctcaaa
     3541 tgacagtcct gtctgtgaca aattgccctt aaccctgtga caaattgccc tcagaagaag
     3601 ctgttttttc acaaagttat ccctgcttat tgactctttt ttatttagtg tgacaatcta
     3661 aaaacttgtc acacttcaca tggatctgtc atggcggaaa cagcggttat caatcacaag
     3721 aaacgtaaaa atagcccgcg aatcgtccag tcaaacgacc tcactgaggc ggcatatagt
     3781 ctctcccggg atcaaaaacg tatgctgtat ctgttcgttg accagatcag aaaatctgat
     3841 ggcaccctac aggaacatga cggtatctgc gagatccatg ttgctaaata tgctgaaata
     3901 ttcggattga cctctgcgga agccagtaag gatatacggc aggcattgaa gagtttcgcg
     3961 gggaaggaag tggtttttta tcgccctgaa gaggatgccg gcgatgaaaa aggctatgaa
     4021 tcttttcctt ggtttatcaa acgtgcgcac agtccatcca gagggcttta cagtgtacat
     4081 atcaacccat atctcattcc cttctttatc gggttacaga accggtttac gcagtttcgg
     4141 cttagtgaaa caaaagaaat caccaatccg tatgccatgc gtttatacga atccctgtgt
     4201 cagtatcgta agccggatgg ctcaggcatc gtgtctctga aaatcgactg gatcatagag
     4261 cgttaccagc tgcctcaaag ttaccagcgt atgcctgact tccgccgccg cttcctgcag
     4321 gtctgtgtta atgagatcaa cagcagaact ccaatgcgcc tctcatacat tgagaaaaag
     4381 aaaggccgcc agacgactca tatcgtattt tccttccgcg atatcacttc catgacgaca
     4441 ggatagtctg agggttatct gtcacagatt tgagggtggt tcgtcacatt tgttctgacc
     4501 tactgagggt aatttgtcac agttttgctg tttccttcag cctgcatgga ttttctcata
     4561 ctttttgaac tgtaattttt aaggaagcca aatttgaggg cagtttgtca cagttgattt
     4621 ccttctcttt cccttcgtca tgtgacctga tatcgggggt tagttcgtca tcattgatga
     4681 gggttgatta tcacagttta ttactctgaa ttggctatcc gcgtgtgtac ctctacctgg
     4741 agtttttccc acggtggata tttcttcttg cgctgagcgt aagagctatc tgacagaaca
     4801 gttcttcttt gcttcctcgc cagttcgctc gctatgctcg gttacacggc tgcggcgagc
     4861 gctagtgata ataagtgact gaggtatgtg gtgttcttat ctccttttgt agtgttgctc
     4921 ttattttaaa caactttgcg gttttttgat gactttgcga ttttgttgtt gctttgcagt
     4981 aaattgcaag atttaataaa aaaacgcaaa gcaatgatta aaggatgttc agaatgaaac
     5041 tcatggaaac acttaaccag tgcataaacg ctggtcatga aatgacgaag gctatcgcca
     5101 ttgcacagtt taatgatgac agcccggaag cgaggaaaat aacccggcgc tggagaatag
     5161 gtgaagcagc ggatttagtt ggggtttctt ctcaggctat cagagatgcc gagaaagcag
     5221 ggcgactacc gcacccggat atggaaattc gaggacgggt tgagcaacgt gttggttata
     5281 caattgaaca aattaatcat atgcgtgatg tgtttggtac gcgattgcga cgtgctgagg
     5341 acgtatttcc accggtgatc ggggttgctg cccataaagg tggcgtttac aaaacctcag
     5401 tttctgttca tcttgctcag gatctggctc tgaaggggct acgtgttttg ctcgtggaag
     5461 gtaacgaccc ccagggaaca gcctcaatgt atcacggatg ggtaccagat cttcatattc
     5521 atgcagaaga tactctcctg cctttctatc ttggggaaaa ggacgatgtc acttatgcaa
     5581 taaagcccac ttgctggccg gggcttgaca ttattccttc ctgtctggct ctgcaccgta
     5641 ttgaaactga gttaatgggc aaatttgatg aaggtaaact gcccaccgat ccacacctga
     5701 tgctccgact ggccattgaa actgttgctc atgactatga tgtcatagtt attgacagcg
     5761 cgcctaacct gggtatcggc acgattaatg tcgtatgtgc tgctgatgtg ctgattgttc
     5821 ccacgcctgc tgagttgttt gactacacct ccgcactgca gtttttcgat atgcttcgtg
     5881 atctgctcaa gaacgttgat cttaaagggt tcgagcctga tgtacgtatt ttgcttacca
     5941 aatacagcaa tagtaatggc tctcagtccc cgtggatgga ggagcaaatt cgggatgcct
     6001 ggggaagcat ggttctaaaa aatgttgtac gtgaaacgga tgaagttggt aaaggtcaga
     6061 tccggatgag aactgttttt gaacaggcca ttgatcaacg cagttcaact ggtgcctgga
     6121 gaaatgctct ttctatttgg gaacctgtct gcaatgaaat tttcgatcgt ctgattaaac
     6181 cacgctggga gattagataa tgaagcgtgc gcctgttatt ccaaaacata cgctcaatac
     6241 tcaaccggtt gaagatactt cgttatcgac accagctgcc ccgatggtgg attcgttaat
     6301 tgcgcgcgta ggagtaatgg ctcgcggtaa tgccattact ttgcctgtat gtggtcggga
     6361 tgtgaagttt actcttgaag tgctccgggg tgatagtgtt gagaaaacct ctcgggtatg
     6421 gtcaggtaat gaacgtgacc aggagctgct tactgaggac gcactggatg atctcatccc
     6481 ttcttttcta ctgactggtc aacagacacc ggcgttcggt cgaagagtat ctggtgtcat
     6541 agaaattgcc gatgggagtc gccgtcgtaa agctgctgca cttaccgaaa gtgattatcg
     6601 tgttctggtt ggcgagctgg atgatgagca gatggctgca ttatccagat tgggtaacga
     6661 ttatcgccca acaagtgctt atgaacgtgg tcagcgttat gcaagccgat tgcagaatga
     6721 atttgctgga aatatttctg cgctggctga tgcggaaaat atttcacgta agattattac
     6781 ccgctgtatc aacaccgcca aattgcctaa atcagttgtt gctctttttt ctcaccccgg
     6841 tgaactatct gcccggtcag gtgatgcact tcaaaaagcc tttacagata aagaggaatt
     6901 acttaagcag caggcatcta accttcatga gcagaaaaaa gctggggtga tatttgaagc
     6961 tgaagaagtt atcactcttt taacttctgt gcttaaaacg tcatctgcat caagaactag
     7021 tttaagctca cgacatcagt ttgctcctgg agcgacagta ttgtataagg gcgataaaat
     7081 ggtgcttaac ctggacaggt cacgtgttcc aactgagtgt atagagaaaa ttgaggccat
     7141 tcttaaggaa cttgaaaagc cagcaccctg atgcgaccac gttttagtct acgtttatct
     7201 gtctttactt aatgtccttt gttacaggcc agaaagcata actggcctga atattctctc
     7261 tgggcccact gttccacttg tatcgtcggt ctgataatca gactgggacc acggtcccac
     7321 tcgtatcgtc ggtctgatta ttagtctggg accacggtcc cactcgtatc gtcggtctga
     7381 ttattagtct gggaccacgg tcccactcgt atcgtcggtc tgataatcag actgggacca
     7441 cggtcccact cgtatcgtcg gtctgattat tagtctggga ccatggtccc actcgtatcg
     7501 tcggtctgat tattagtctg ggaccacggt cccactcgta tcgtcggtct gattattagt
     7561 ctggaaccac ggtcccactc gtatcgtcgg tctgattatt agtctgggac cacggtccca
     7621 ctcgtatcgt cggtctgatt attagtctgg gaccacgatc ccactcgtgt tgtcggtctg
     7681 attatcggtc tgggaccacg gtcccacttg tattgtcgat cagactatca gcgtgagact
     7741 acgattccat caatgcctgt caagggcaag tattgacatg tcgtcgtaac ctgtagaacg
     7801 gagtaacctc ggtgtgcggt tgtatgcctg ctgtggattg ctgctgtgtc ctgcttatcc
     7861 acaacatttt gcgcacggtt atgtggacaa aatacctggt tacccaggcc gtgccggcac
     7921 gttaaccggg ctgcatccga tgcaagtgtg tcgctgtcga cgcaagcggc cgca
//
