LOCUS       Exported                7309 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Broad host-range plasmid with toxin-antitoxin system for maintenance
            in the absence of antibiotic selection, and constitutive promoter.
ACCESSION   .
VERSION     .
KEYWORDS    pBBR5pemIKpKan
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7309)
  AUTHORS   Burbank L, Wei W
  TITLE     Broad-Host-Range Plasmids for Constitutive and Inducible Gene 
            Expression in the Absence of Antibiotic Selection.
  JOURNAL   Microbiol Resour Announc. 2019 Sep 5;8(36). pii: 8/36/e00769-19. 
            doi: 10.1128/MRA.00769-19.
  PUBMED    31488530
REFERENCE   2  (bases 1 to 7309)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7309
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      66..837
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             838..1500
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     primer_bind     2194..2216
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2208..2225
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2209..2225
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2235..2254
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2235..2253
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     2406..2422
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2406..2422
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    2431..2555
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        2580..2610
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2664..3323
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2730..2749)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             3665..3970
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3849..3868
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4011..4135)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(4874..4890)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4875..4891)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(4920..4940)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4920..4938)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4959..4975)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4959..4975)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4972..4994)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4983..4999
                     /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(5007..5037)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5052..5073
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5343..5371
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             5560..6093
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
ORIGIN
        1 ggcttcgccc tgcgcggtcg ctgcgctccc ttgccagccc gtggatatgt ggacgatggc
       61 cgcgagcggc caccggctgg ctcgcttcgc tcggcccgtg gacaaccctg ctggacaagc
      121 tgatggacag gctgcgcctg cccacgagct tgaccacagg gattgcccac cggctaccca
      181 gccttcgacc acatacccac cggctccaac tgcgcggcct gcggccttgc cccatcaatt
      241 tttttaattt tctctgggga aaagcctccg gcctgcggcc tgcgcgcttc gcttgccggt
      301 tggacaccaa gtggaaggcg ggtcaaggct cgcgcagcga ccgcgcagcg gcttggcctt
      361 gacgcgcctg gaacgaccca agcctatgcg agtgggggca gtcgaagggc gaagcccgcc
      421 cgcctgcccc ccgagcctca cggcggcgag tgcgggggtt ccaagggggc agcgccacct
      481 tgggcaaggc cgaaggccgc gcagtcgatc aacaagcccc ggaggggcca ctttttgccg
      541 gagggggagc cgcgccgaag gcgtggggga accccgcagg ggtgcccttc tttgggcacc
      601 aaagaactag atatagggcg aaatgcgaaa gacttaaaaa tcaacaactt aaaaaagggg
      661 ggtacgcaac agctcattgc ggcacccccc gcaatagctc attgcgtagg ttaaagaaaa
      721 tctgtaattg actgccactt ttacgcaacg cataattgtt gtcgcgctgc cgaaaagttg
      781 cagctgattg cgcatggtgc cgcaaccgtg cggcacccct accgcatgga gataagcatg
      841 gccacgcagt ccagagaaat cggcattcaa gccaagaaca agcccggtca ctgggtgcaa
      901 acggaacgca aagcgcatga ggcgtgggcc gggcttattg cgaggaaacc cacggcggca
      961 atgctgctgc atcacctcgt ggcgcagatg ggccaccaga acgccgtggt ggtcagccag
     1021 aagacacttt ccaagctcat cggacgttct ttgcggacgg tccaatacgc agtcaaggac
     1081 ttggtggccg agcgctggat ctccgtcgtg aagctcaacg gccccggcac cgtgtcggcc
     1141 tacgtggtca atgaccgcgt ggcgtggggc cagccccgcg accagttgcg cctgtcggtg
     1201 ttcagtgccg ccgtggtggt tgatcacgac gaccaggacg aatcgctgtt ggggcatggc
     1261 gacctgcgcc gcatcccgac cctgtatccg ggcgagcagc aactaccgac cggccccggc
     1321 gaggagccgc ccagccagcc cggcattccg ggcatggaac cagacctgcc agccttgacc
     1381 gaaacggagg aatgggaacg gcgcgggcag cagcgcctgc cgatgcccga tgagccgtgt
     1441 tttctggacg atggcgagcc gttggagccg ccgacacggg tcacgctgcc gcgccggtag
     1501 cacttgggtt gcgcagcaac ccgtaagtgc gctgttccag actatcggct gtagccgcct
     1561 cgccgcccta taccttgtct gcctccccgc gttgcgtcgc ggtgcatgga gccgggccac
     1621 ctcgacctga atggaagccg gcggcacctc gctaacggat tcaccgtttt tatcaggctc
     1681 tgggaggcag aataaatgat catatcgtca attattacct ccacggggag agcctgagca
     1741 aactggcctc aggcatttga gaagcacacg gtcacactgc ttccggtagt caataaaccg
     1801 gtaaaccagc aatagacata agcggctatt taacgaccct gccctgaacc gacgaccggg
     1861 tcgaatttgc tttcgaattt ctgccattca tccgcttatt atcacttatt caggcgtagc
     1921 aaccaggcgt ttaagggcac caataactgc cttaaaaaaa ttacgccccg ccctgccact
     1981 catcgcagta cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     2041 tttaacaaaa tattaacgct tacaatttcc attcgccatt caggctgcgc aactgttggg
     2101 aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg
     2161 caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg
     2221 ccagtgagcg cgcgtaatac gactcactat agggcgaatt ggagctcccg gaattgccag
     2281 ctggggcgcc ctctggtaag gttgggaagc cctgcaaagt aaactggatg gctttcttgc
     2341 cgccaaggat ctgatggcgc aggggatcaa gatctgatca agagacagga tgaggatcgt
     2401 ttcgctctag aactagtgga tcccccatca acaagtttgt acaaaaaagc tgaacgagaa
     2461 acgtaaaatg atataaatat caatatatta aattagattt tgcataaaaa acagactaca
     2521 taatactgta aaacacaaca tatccagtca ctatggcggc cgcattaggc accccaggct
     2581 ttacacttta tgcttccggc tcgtataatg tgtggatttt gagttaggat ccgtcgagat
     2641 tttcaggagc taaggaagct aaaatggaga aaaaaatcac tggatatacc accgttgata
     2701 tatcccaatg gcatcgtaaa gaacattttg aggcatttca gtcagttgct caatgtacct
     2761 ataaccagac cgttcagctg gatattacgg cctttttaaa gaccgtaaag aaaaataagc
     2821 acaagtttta tccggccttt attcacattc ttgcccgcct gatgaatgct catccggaat
     2881 tccgtatggc aatgaaagac ggtgagctgg tgatatggga tagtgttcac ccttgttaca
     2941 ccgttttcca tgagcaaact gaaacgtttt catcgctctg gagtgaatac cacgacgatt
     3001 tccggcagtt tctacacata tattcgcaag atgtggcgtg ttacggtgaa aacctggcct
     3061 atttccctaa agggtttatt gagaatatgt ttttcgtctc agccaatccc tgggtgagtt
     3121 tcaccagttt tgatttaaac gtggccaata tggacaactt cttcgccccc gttttcacca
     3181 tgggcaaata ttatacgcaa ggcgacaagg tgctgatgcc gctggcgatt caggttcatc
     3241 atgccgtttg tgatggcttc catgtcggca gaatgcttaa tgaattacaa cagtactgcg
     3301 atgagtggca gggcggggcg taaagatctg gatccggctt actaaaagcc agataacagt
     3361 atgcgtattt gcgcgctgat ttttgcggta taagaatata tactgatatg tatacccgaa
     3421 gtatgtcaaa aagaggtatg ctatgaagca gcgtattaca gtgacagttg acagcgacag
     3481 ctatcagttg ctcaaggcat atatgatgtc aatatctccg gtctggtaag cacaaccatg
     3541 cagaatgaag cccgtcgtct gcgtgccgaa cgctggaaag cggaaaatca ggaagggatg
     3601 gctgaggtcg cccggtttat tgaaatgaac ggctcttttg ctgacgagaa caggggctgg
     3661 tgaaatgcag tttaaggttt acacctataa aagagagagc cgttatcgtc tgtttgtgga
     3721 tgtacagagt gatattattg acacgcccgg gcgacggatg gtgatccccc tggccagtgc
     3781 acgtctgctg tcagataaag tctcccgtga actttacccg gtggtgcata tcggggatga
     3841 aagctggcgc atgatgacca ccgatatggc cagtgtgccg gtctccgtta tcggggaaga
     3901 agtggctgat ctcagccacc gcgaaaatga catcaaaaac gccattaacc tgatgttctg
     3961 gggaatataa atgtcaggct cccttataca cagccagtct gcaggtcgac catagtgact
     4021 ggatatgttg tgttttacag tattatgtag tctgtttttt atgcaaaatc taatttaata
     4081 tattgatatt tatatcattt tacgtttctc gttcagcttt cttgtacaaa gtggttgatg
     4141 ggctgcagga attcgatttt cgctataagc gtataaatgg aggaattata ttatacttta
     4201 atataactta acgaaagaag cccatcatgc acaccacaaa ccttcgtaaa gtcggtggat
     4261 cgatcatgct ggccgttccc cccgctttcc ttgatcagct tcatcttgag gtcggcgcaa
     4321 cggtcggctt agccgtcaca gacggtcgtc tggtcattga gccagtccta catcctcagt
     4381 acacgctgga tcaactgctg gccgaagcgg aagcctccgg ggcttacccg ttgccgcccg
     4441 aggaacgtga atgggtcgat gctccagcag taggacgcga actgatatga aacgcggcga
     4501 tgtttacatg gtcgatctgg agccaaccgc cggacacgaa caacgcggtc accgtccggt
     4561 ggttgttatc tccagtgagc gattcaatcg gcttacctct tgtccggtga tccttccaat
     4621 cacaaacggc ggcgagttcg cctcccgcct cgggttcgct gtggaattac taggtacgat
     4681 tacaacgggc gtcgttcgat gtgatcaacc acgtgcgctc gacttactcg ctcgcaatgc
     4741 ccgcaaagtg gaaagcctac caccaaacat cttggccgat gtcttggcaa aagctgtcac
     4801 catcttccag taatttttgg ggcaagttca ggccactgca accaatcact agtgaattcg
     4861 atatcaagct tatcgatacc gtcgacctcg agggggggcc cggtacccag cttttgttcc
     4921 ctttagtgag ggttaattgc gcgcttggcg taatcatggt catagctgtt tcctgtgtga
     4981 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     5041 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     5101 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     5161 ggtttgcgta ttgggcgcat gcataaaaac tgttgtaatt cattaagcat tctgccgaca
     5221 tggaagccat cacaaacggc atgatgaacc tgaatcgcca gcggcatcag caccttgtcg
     5281 ccttgcgtat aatatttgcc catggacgca caccgtggaa acggatgaag gcacgaaccc
     5341 agttgacata agcctgttcg gttcgtaaac tgtaatgcaa gtagcgtatg cgctcacgca
     5401 actggtccag aaccttgacc gaacgcagcg gtggtaacgg cgcagtggcg gttttcatgg
     5461 cttgttatga ctgttttttt gtacagtcta tgcctcgggc atccaagcag caagcgcgtt
     5521 acgccgtggg tcgatgtttg atgttatgga gcagcaacga tgttacgcag cagcaacgat
     5581 gttacgcagc agggcagtcg ccctaaaaca aagttaggtg gctcaagtat gggcatcatt
     5641 cgcacatgta ggctcggccc tgaccaagtc aaatccatgc gggctgctct tgatcttttc
     5701 ggtcgtgagt tcggagacgt agccacctac tcccaacatc agccggactc cgattacctc
     5761 gggaacttgc tccgtagtaa gacattcatc gcgcttgctg ccttcgacca agaagcggtt
     5821 gttggcgctc tcgcggctta cgttctgccc aagtttgagc agccgcgtag tgagatctat
     5881 atctatgatc tcgcagtctc cggagagcac cggaggcagg gcattgccac cgcgctcatc
     5941 aatctcctca agcatgaggc caacgcgctt ggtgcttatg tgatctacgt gcaagcagat
     6001 tacggtgacg atcccgcagt ggctctctat acaaagttgg gcatacggga agaagtgatg
     6061 cactttgata tcgacccaag taccgccacc taacaattcg ttcaagccga gatcggcttc
     6121 ccggccgcgg agttgttcgg taaattgtca caacgccgca ggtggcactt ttcggggaaa
     6181 tgtgcgcgcc cgcgttcctg ctggcgctgg gcctgtttct ggcgctggac ttcccgctgt
     6241 tccgtcagca gcttttcgcc cacggccttg atgatcgcgg cggccttggc ctgcatatcc
     6301 cgattcaacg gccccagggc gtccagaacg ggcttcaggc gctcccgaag gtctcgggcc
     6361 gtctcttggg cttgatcggc cttcttgcgc atctcacgcg ctcctgcggc ggcctgtagg
     6421 gcaggctcat acccctgccg aaccgctttt gtcagccggt cggccacggc ttccggcgtc
     6481 tcaacgcgct ttgagattcc cagcttttcg gccaatccct gcggtgcata ggcgcgtggc
     6541 tcgaccgctt gcgggctgat ggtgacgtgg cccactggtg gccgctccag ggcctcgtag
     6601 aacgcctgaa tgcgcgtgtg acgtgccttg ctgccctcga tgccccgttg cagccctaga
     6661 tcggccacag cggccgcaaa cgtggtctgg tcgcgggtca tctgcgcttt gttgccgatg
     6721 aactccttgg ccgacagcct gccgtcctgc gtcagcggca ccacgaacgc ggtcatgtgc
     6781 gggctggttt cgtcacggtg gatgctggcc gtcacgatgc gatccgcccc gtacttgtcc
     6841 gccagccact tgtgcgcctt ctcgaagaac gccgcctgct gttcttggct ggccgacttc
     6901 caccattccg ggctggccgt catgacgtac tcgaccgcca acacagcgtc cttgcgccgc
     6961 ttctctggca gcaactcgcg cagtcggccc atcgcttcat cggtgctgct ggccgcccag
     7021 tgctcgttct ctggcgtcct gctggcgtca gcgttgggcg tctcgcgctc gcggtaggcg
     7081 tgcttgagac tggccgccac gttgcccatt ttcgccagct tcttgcatcg catgatcgcg
     7141 tatgccgcca tgcctgcccc tcccttttgg tgtccaaccg gctcgacggg ggcagcgcaa
     7201 ggcggtgcct ccggcgggcc actcaatgct tgagtatact cactagactt tgcttcgcaa
     7261 agtcgtgacc gcctacggcg gctgcggcgc cctacgggct tgctctccg
//
