LOCUS       sequence-269060-        3200 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3200)
  AUTHORS   Azubuike CC, Gatehouse AMR, Howard TP
  TITLE     pCAT vectors overcome inefficient electroporation of Cupriavidus 
            necator H16.
  JOURNAL   N Biotechnol. 2021 Nov 25;65:20-30. doi: 10.1016/j.nbt.2021.07.003. 
            Epub 2021 Jul 29.
  PUBMED    34333160
REFERENCE   2  (bases 1 to 3200)
  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: "N 
            Biotechnol."; date: "2021-11-25"; volume: "65:20-30. doi"; pages: " 
            10.1016/j.nbt.2021.07.003. Epub 2021 Jul 29"
COMMENT     Sequence Label: pCAT201
FEATURES             Location/Qualifiers
     source          1..3200
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(52..57,58..75,76..82)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    90..106
                     /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     95..117
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     114..130
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     114..130
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             126..491
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWVPGPPSRSTVSISLISNSCSPGDPLV
                     LERPPPRWSSNSPYSESYYARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEAR
                     TDRPSQQLRSLNGEWKL"
     primer_bind     149..169
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        151..169
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    182..289
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     198..214
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     199..215
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(249..265)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(249..265)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(297..316)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(298..316)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(326..343)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(326..342)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(335..357)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(665..1327)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      1328..2099
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     primer_bind     complement(2335..2354)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2346..3161
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             2346..3161
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2419..2438)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 acgctgactt gacgggacgg cgcgccgctt cactcattag gcaccccagg ctttacactt
       61 tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa
      121 cagctatgac catgattacg ccaagcgcgc aattaaccct cactaaaggg aacaaaagct
      181 gggtaccggg ccccccctcg aggtcgacgg tatcgataag cttgatatcg aattcctgca
      241 gcccggggga tccactagtt ctagagcggc cgccaccgcg gtggagctcc aattcgccct
      301 atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt acaacgtcgt gactgggaaa
      361 accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta
      421 atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat
      481 ggaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
      541 attttttaac caataggccg actgcgatga gttaattaag ttccagacaa ggtatagggc
      601 ggcgaggcgg ctacagccga tagtctggaa cagcgcactt acgggttgct gcgcaaccca
      661 agtgctaccg gcgcggcagc gtgacccgtg tcggcggctc caacggctcg ccatcgtcca
      721 gaaaacacgg ctcatcgggc atcggcaggc gctgctgccc gcgccgttcc cattcctccg
      781 tttcggtcaa ggctggcagg tctggttcca tgcccggaat gccgggctgg ctgggcggct
      841 cctcgccggg gccggtcggt agttgctgct cgcccggata cagggtcggg atgcggcgca
      901 ggtcgccatg ccccaacagc gattcgtcct ggtcgtcgtg atcaaccacc acggcggcac
      961 tgaacaccga caggcgcaac tggtcgcggg gctggcccca cgccacgcgg tcattgacca
     1021 cgtaggccga cacggtgccg gggccgttga gcttcacgac ggagatccag cgctcggcca
     1081 ccaagtcctt gactgcgtat tggaccgtcc gcaaagaacg tccgatgagc ttggaaagtg
     1141 tcttctggct gaccaccacg gcgttctggt ggcccatctg cgccacgagg tgatgcagca
     1201 gcattgccgc cgtgggtttc ctcgcaataa gcccggccca cgcctcatgc gctttgcgtt
     1261 ccgtttgcac ccagtgaccg ggcttgttct tggcttgaat gccgatttct ctggactgcg
     1321 tggccatgct tatctccatg cggtaggggt gccgcacggt tgcggcacca tgcgcaatca
     1381 gctgcaactt ttcggcagcg cgacaacaat tatgcgttgc gtaaaagtgg cagtcaatta
     1441 cagattttct ttaacctacg caatgagcta ttgcgggggg tgccgcaatg agctgttgcg
     1501 tacccccctt ttttaagttg ttgattttta agtctttcgc atttcgccct atatctagtt
     1561 ctttggtgcc caaagaaggg cacccctgcg gggttccccc acgccttcgg cgcggctccc
     1621 cctccggcaa aaagtggccc ctccggggct tgttgatcga ctgcgcggcc ttcggccttg
     1681 cccaaggtgg cgctgccccc ttggaacccc cgcactcgcc gccgtgaggc tcggggggca
     1741 ggcgggcggg cttcgccctt cgactgcccc cactcgcata ggcttgggtc gttccaggcg
     1801 cgtcaaggcc aagccgctgc gcggtcgctg cgcgagcctt gacccgcctt ccacttggtg
     1861 tccaaccggc aagcgaagcg cgcaggccgc aggccggagg cttttcccca gagaaaatta
     1921 aaaaaattga tggggcaagg ccgcaggccg cgcagttgga gccggtgggt atgtggtcga
     1981 aggctgggta gccggtgggc aatccctgtg gtcaagctcg tgggcaggcg cagcctgtcc
     2041 atcagcttgt ccagcagggt tgtccacggg ccgagcgaag cgagccagcc ggtggccgct
     2101 cgcggccatc gtccacatat ccacgggctg gcaagggagc gcagcgaccg cgcagggcga
     2161 agcccggaga gcaagcccgt agggcgccgc agccgccgta ggcggtcacg actttgcgaa
     2221 gggccggcca cctggaaagc cacgttgtgt ctcaaaatct ctgatgttac attgcacaag
     2281 ataaaaatat atcatcatga acaataaaac tgtctgctta cataaacagt aatacaaggg
     2341 gtgttatgag ccatattcaa cgggaaacgt cttgctcgag gccgcgatta aattccaaca
     2401 tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga
     2461 caatctatcg attgtatggg aagcccgatg cgccagagtt gtttctgaaa catggcaaag
     2521 gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg acggaattta
     2581 tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg ttactcacca
     2641 ctgcgatccc cgggaaaaca gcattccagg tattagaaga atatcctgat tcaggtgaaa
     2701 atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct gtttgtaatt
     2761 gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga atgaataacg
     2821 gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct
     2881 ggaaagaaat gcataagctt ttgccattct caccggattc agtcgtcact catggtgatt
     2941 tctcacttga taaccttatt tttgacgagg ggaaattaat aggttgtatt gatgttggac
     3001 gagtcggaat cgcagaccga taccaggatc ttgccatcct atggaactgc ctcggtgagt
     3061 tttctccttc attacagaaa cggctttttc aaaaatatgg tattgataat cctgatatga
     3121 ataaattgca gtttcatttg atgctcgatg agtttttcta atcagaattg gttaattggt
     3181 tgtaacactg gcagagcatt
//
