LOCUS       sequence-388027-        3211 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 1 destination vector containing pBBR1 origin of replication 
            and CIDAR AE scars.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3211)
  AUTHORS   Keating KW, Young EM
  TITLE     Systematic Part Transfer by Extending a Modular Toolkit to Diverse 
            Bacteria.
  JOURNAL   ACS Synth Biol. 2023 Jun 9. doi: 10.1021/acssynbio.3c00104.
  PUBMED    37294017
REFERENCE   2  (bases 1 to 3211)
  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: "ACS Synth 
            Biol. 2023 Jun 9. doi: 10.1021/acssynbio.3c00104."
COMMENT     Sequence Label: pBBR1_DVK_AE
FEATURES             Location/Qualifiers
     source          1..3211
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     198..220
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     212..229
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     213..229
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    230..286
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(299..315)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(299..315)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(312..334)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    323..339
                     /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(347..353,354..371,372..377))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    392..413
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      642..1413
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             1414..2076
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     oriT            2087..2195
                     /note="incP origin of transfer"
     primer_bind     complement(2285..2304)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2296..3111
                     /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             2296..3111
                     /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(2369..2388)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(join(3146..3211,1..303))
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCRSLSMSS
                     TRSDRVLGSDAQWNEKSI"
ORIGIN
        1 ggagagagac ctgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata
       61 ccgcatcagg cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
      121 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
      181 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg aattcgagct
      241 cggtacccgg ggatcctcta gagtcgacct gcaggcatgc aagcttggcg taatcatggt
      301 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
      361 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
      421 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
      481 gccaacgcgc gggggtctct gcttatgtct tcagcagtag gacaaatccg ccgccctaga
      541 cagctgggcg cgccccccct acgggcttgc tctccgggct tcgccctgcg cggtcgctgc
      601 gctcccttgc cagcccgtgg atatgtggac gatggccgcg agcggccacc ggctggctcg
      661 cttcgctcgg cccgtggaca accctgctgg acaagctgat ggacaggctg cgcctgccca
      721 cgagcttgac cacagggatt gcccaccggc tacccagcct tcgaccacat acccaccggc
      781 tccaactgcg cggcctgcgg ccttgcccca tcaatttttt taattttctc tggggaaaag
      841 cctccggcct gcggcctgcg cgcttcgctt gccggttgga caccaagtgg aaggcgggtc
      901 aaggctcgcg cagcgaccgc gcagcggctt ggccttgacg cgcctggaac gacccaagcc
      961 tatgcgagtg ggggcagtcg aagggcgaag cccgcccgcc tgccccccga gcctcacggc
     1021 ggcgagtgcg ggggttccaa gggggcagcg ccaccttggg caaggccgaa ggccgcgcag
     1081 tcgatcaaca agccccggag gggccacttt ttgccggagg gggagccgcg ccgaaggcgt
     1141 gggggaaccc cgcaggggtg cccttctttg ggcaccaaag aactagatat agggcgaaat
     1201 gcgaaagact taaaaatcaa caacttaaaa aaggggggta cgcaacagct cattgcggca
     1261 ccccccgcaa tagctcattg cgtaggttaa agaaaatctg taattgactg ccacttttac
     1321 gcaacgcata attgttgtcg cgctgccgaa aagttgcagc tgattgcgca tggtgccgca
     1381 accgtgcggc acccctaccg catggagata agcatggcca cgcagtccag agaaatcggc
     1441 attcaagcca agaacaagcc cggtcactgg gtgcaaacgg aacgcaaagc gcatgaggcg
     1501 tgggccgggc ttattgcgag gaaacccacg gcggcaatgc tgctgcatca cctcgtggcg
     1561 cagatgggcc accagaacgc cgtggtggtc agccagaaaa cactttccaa gctcatcgga
     1621 cgttctttgc ggacggtcca atacgcagtc aaggacttgg tggccgagcg ctggatctcc
     1681 gtcgtgaagc tcaacggccc cggcaccgtg tcggcctacg tggtcaatga ccgcgtggcg
     1741 tggggccagc cccgcgacca gttgcgcctg tcggtgttca gtgccgccgt ggtggttgat
     1801 cacgacgacc aggacgaatc gctgttgggg catggcgacc tgcgccgcat cccgaccctg
     1861 tatccgggcg agcagcaact accgaccggc cccggcgagg agccgcccag ccagcccggc
     1921 attccgggca tggaaccaga cctgccagcc ttgaccgaaa cggaggaatg ggaacggcgc
     1981 gggcagcagc gcctgccgat gcccgatgag ccgtgttttc tggacgatgg cgagccgttg
     2041 gagccgccga cacgggtaac gctgccgcgc cggtagggcc ggcctacggc cagcctcgca
     2101 gagcaggatt cccgttgagc accgccaggt gcgaataagg gacagtgaag aaggaacacc
     2161 cgctcgcggg tgggcctact tcacctatcc tgcccttgtg tctcaaaatc tctgatgtac
     2221 attgcacaag ataaaaatat atcatcatga acaataaaac tgtctgctta cataaacagt
     2281 aatacaaggg gtgttatgag ccatattcaa cgggaaacgt cttgctccag gccgcgatta
     2341 aattccaaca tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa
     2401 tcaggtgcga caatctatcg attgtatggg aagcccgatg cgccagagtt gtttctgaaa
     2461 catggcaaag gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg
     2521 acggaattta tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg
     2581 ttactcacca ctgcgatccc cgggaaaaca gcattccagg tattagaaga atatcctgat
     2641 tcaggtgaaa atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct
     2701 gtttgtaatt gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga
     2761 atgaataacg gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt
     2821 gaacaagtct ggaaagaaat gcataagctt ttgccattct caccggattc agtcgtcact
     2881 catggtgatt tctcacttga taaccttatt tttgacgagg ggaaattaat aggttgtatt
     2941 gatgttggac gagtcggaat cgcagaccga taccaggatc ttgccatcct atggaactgc
     3001 ctcggtgagt tttctccttc attacagaaa cggctttttc aaaaatatgg tattgataat
     3061 cctgatatga ataaattgca gtttcatttg atgctcgatg agtttttcta aggtaactgt
     3121 cagaccaagt ttactcatat atactttaga ttgatttttc gttccactga gcgtcagacc
     3181 ccagaactct gtcagaccga gtggaagaca t
//
