LOCUS       sequence-220098-        7504 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescence microscopy.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7504)
  AUTHORS   Lefebre MD, Valvano MA
  TITLE     Construction and evaluation of plasmid vectors optimized for 
            constitutive and regulated gene expression in Burkholderia cepacia 
            complex isolates.
  JOURNAL   Appl Environ Microbiol. 2002 Dec;68(12):5956-64.
  PUBMED    12450816
REFERENCE   2  (bases 1 to 7504)
  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: "Appl 
            Environ Microbiol."; date: "2002-12"; volume: "68(12)"; pages: 
            "5956-64"
COMMENT     Sequence Label: pMLBAD-eGFP
FEATURES             Location/Qualifiers
     source          1..7504
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..285
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     208..227
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             join(320..322,323..325,326..1039)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(320..322,323..325,326..1036)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(365..386)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(626..645)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     973..994
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1129..1146)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1299..1385
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1477..1504
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     2272..2291
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(2358..2460)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(2577..2595)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     rep_origin      3853..4624
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             4625..5287
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     promoter        join(5787..5792,5793..5809,5810..5815)
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             6141..6377
                     /codon_start=1
                     /gene="dhfrIIb from plasmid R388"
                     /product="E. coli plasmid-associated dihydrofolate 
                     reductase"
                     /label=TpR
                     /note="confers resistance to trimethoprim"
                     /translation="MGQSSDEANAPVAGQFALPLSATFGLGDRVRKKSGAAWQGQVVGW
                     YCTKLTPEGYAVESESHPGSVQIYPVAALERVA"
     primer_bind     6401..6420
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(6600..7478)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct
       61 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
      121 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
      181 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
      241 atcctacctg acgcttttta tcgcaactct ctactgtttc tccatacccg tttttttggg
      301 ctagcaggag gaattcacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
      361 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
      421 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
      481 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
      541 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
      601 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
      661 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
      721 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
      781 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga
      841 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
      901 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga
      961 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
     1021 ggacgagctg tacaagtaaa gcggccgcga attcgatcgg ggatcctcta gagtcgacct
     1081 gcaggcatgc aagcttggct gttttggcgg atgagagaag attttcagcc tgatacagat
     1141 taaatcagaa cgcagaagcg gtctgataaa acagaatttg cctggcggca gtagcgcggt
     1201 ggtcccacct gaccccatgc cgaactcaga agtgaaacgc cgtagcgccg atggtagtgt
     1261 ggggtctccc catgcgagag tagggaactg ccaggcatca aataaaacga aaggctcagt
     1321 cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc ctgagtagga
     1381 caaatccgcc gggagcggat ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag
     1441 gacgcccgcc ataaactgcc aggcatcaaa ttaagcagaa ggccatcctg acggatggcc
     1501 tttttgcgtt tctacaaact cttccactca ctacacagag ccatttaaac aacatcccct
     1561 ccccctttcc accgcgtcag acgcccgtag cagcccgcta cgggcttttt catgccctgc
     1621 cctagcgtcc aagcctcacg gccgcgctcg gcctctctgg cggccttctg gcgctgaggt
     1681 ctgcctcgtg aagaaggtgt tgctgactca taccaggcct gaatcgcccc atcatccagc
     1741 cagaaagtga gggagccacg gttgatgaga gctttgttgt aggtggacca gttggtgatt
     1801 ttgaactttt gctttgccac ggaacggtct gcgttgtcgg gaagatgcgt gatctgatcc
     1861 ttcaactcag caaaagttcg atttattcaa caaagccgcc gtcccgtcaa gtcagcgtaa
     1921 tgctctgcca gtgttacaac caattaacca attctgatta gaaaaactca tcgagcatca
     1981 aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga aaaagccgtt
     2041 tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga tcctggtatc
     2101 ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa
     2161 taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag aatggcaaaa
     2221 gctaaaaagg ccgtaatatc cagctgaacg gtctggttat aggtacattg agcaactgac
     2281 tgaaatgcct caaaatgttc tttacgatgc cattgggata tatcaacggt ggtatatcca
     2341 gtgatttttt tctccatttt agcttcctta gctcctgaaa atctcgataa ctcaaaaaat
     2401 acgcccggta gtgatcttat ttcattatgg tgaaagttgg aacctcttac gtgccgatca
     2461 acgtctcatt ttcgccaaaa gttggcccag ggcttcccgg tatcaacagg gacaccagga
     2521 tttatttatt ctgcgaagtg atcttccgtc acaggtattt attcgaagac gaaagggcct
     2581 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     2641 tggcactttt cggggaaatg tgcgcgcccg cgttcctgct ggcgctgggc ctgtttctgg
     2701 cgctggactt cccgctgttc cgtcagcagc ttttcgccca cggccttgat gatcgcggcg
     2761 gccttggcct gcatatcccg attcaacggc cccagggcgt ccagaacggg cttcaggcgc
     2821 tcccgaaggt ctcgggccgt ctcttgggct tgatcggcct tcttgcgcat ctcacgcgct
     2881 cctgcggcgg cctgtagggc aggctcatac ccctgccgaa ccgcttttgt cagccggtcg
     2941 gccacggctt ccggcgtctc aacgcgcttt gagattccca gcttttcggc caatccctgc
     3001 ggtgcatagg cgcgtggctc gaccgcttgc gggctgatgg tgacgtggcc cactggtggc
     3061 cgctccaggg cctcgtagaa cgcctgaatg cgcgtgtgac gtgccttgct gccctcgatg
     3121 ccccgttgca gccctagatc ggccacagcg gccgcaaacg tggtctggtc gcgggtcatc
     3181 tgcgctttgt tgccgatgaa ctccttggcc gacagcctgc cgtcctgcgt cagcggcacc
     3241 acgaacgcgg tcatgtgcgg gctggtttcg tcacggtgga tgctggccgt cacgatgcga
     3301 tccgccccgt acttgtccgc cagccacttg tgcgccttct cgaagaacgc cgcctgctgt
     3361 tcttggctgg ccgacttcca ccattccggg ctggccgtca tgacgtactc gaccgccaac
     3421 acagcgtcct tgcgccgctt ctctggcagc aactcgcgca gtcggcccat cgcttcatcg
     3481 gtgctgctgg ccgcccagtg ctcgttctct ggcgtcctgc tggcgtcagc gttgggcgtc
     3541 tcgcgctcgc ggtaggcgtg cttgagactg gccgccacgt tgcccatttt cgccagcttc
     3601 ttgcatcgca tgatcgcgta tgccgccatg cctgcccctc ccttttggtg tccaaccggc
     3661 tcgacggggg cagcgcaagg cggtgcctcc ggcgggccac tcaatgcttg agtatactca
     3721 ctagactttg cttcgcaaag tcgtgaccgc ctacggcggc tgcggcgccc tacgggcttg
     3781 ctctccgggc ttcgccctgc gcggtcgctg cgctcccttg ccagcccgtg gatatgtgga
     3841 cgatggccgc gagcggccac cggctggctc gcttcgctcg gcccgtggac aaccctgctg
     3901 gacaagctga tggacaggct gcgcctgccc acgagcttga ccacagggat tgcccaccgg
     3961 ctacccagcc ttcgaccaca tacccaccgg ctccaactgc gcggcctgcg gccttgcccc
     4021 atcaattttt ttaattttct ctggggaaaa gcctccggcc tgcggcctgc gcgcttcgct
     4081 tgccggttgg acaccaagtg gaaggcgggt caaggctcgc gcagcgaccg cgcagcggct
     4141 tggccttgac gcgcctggaa cgacccaagc ctatgcgagt gggggcagtc gaagggcgaa
     4201 gcccgcccgc ctgccccccg agcctcacgg cggcgagtgc gggggttcca agggggcagc
     4261 gccaccttgg gcaaggccga aggccgcgca gtcgatcaac aagccccgga ggggccactt
     4321 tttgccggag ggggagccgc gccgaaggcg tgggggaacc ccgcaggggt gcccttcttt
     4381 gggcaccaaa gaactagata tagggcgaaa tgcgaaagac ttaaaaatca acaacttaaa
     4441 aaaggggggt acgcaacagc tcattgcggc accccccgca atagctcatt gcgtaggtta
     4501 aagaaaatct gtaattgact gccactttta cgcaacgcat aattgttgtc gcgctgccga
     4561 aaagttgcag ctgattgcgc atggtgccgc aaccgtgcgg cacccctacc gcatggagat
     4621 aagcatggcc acgcagtcca gagaaatcgg cattcaagcc aagaacaagc ccggtcactg
     4681 ggtgcaaacg gaacgcaaag cgcatgaggc gtgggccggg cttattgcga ggaaacccac
     4741 ggcggcaatg ctgctgcatc acctcgtggc gcagatgggc caccagaacg ccgtggtggt
     4801 cagccagaag acactttcca agctcatcgg acgttctttg cggacggtcc aatacgcagt
     4861 caaggacttg gtggccgagc gctggatctc cgtcgtgaag ctcaacggcc ccggcaccgt
     4921 gtcggcctac gtggtcaatg accgcgtggc gtggggccag ccccgcgacc agttgcgcct
     4981 gtcggtgttc agtgccgccg tggtggttga tcacgacgac caggacgaat cgctgttggg
     5041 gcatggcgac ctgcgccgca tcccgaccct gtatccgggc gagcagcaac taccgaccgg
     5101 ccccggcgag gagccgccca gccagcccgg cattccgggc atggaaccag acctgccagc
     5161 cttgaccgaa acggaggaat gggaacggcg cgggcagcag cgcctgccga tgcccgatga
     5221 gccgtgtttt ctggacgatg gcgagccgtt ggagccgccg acacgggtca cgctgccgcg
     5281 ccggtagcac ttgggttgcg cagcaacccg taagtgcgct gttccagact atcggctgta
     5341 gccgcctcgc cgccctatac cttgtctgcc tccccgcgtt gcgtcgcggt gcatggagcc
     5401 gggccacctc gacctgaatg gaagccggcg gcacctcgct aacggattca ccgtttttat
     5461 caggctctgg gaggcagaat aaatgatcat atcgtcaatt attacctcca cggggagagc
     5521 ctgagcaaac tggcctcagg catttgagaa gcacacggtc acactgcttc cggtagtcaa
     5581 taaaccggta aaccagcaat agacataagc ggctatttaa cgaccctgcc ctgaaccgac
     5641 gaccgggtcg aatttgcttt cgaatttctg ccattcatcc gcttattatc acttattcag
     5701 gcgtagcacc aggcgtttaa gtcgaccaat aacctacggg gtctgacgct cagtggaacg
     5761 aaatcgatga gctcgcacga acccagttga cataagcctg ttcggttcgt aaactgtaat
     5821 gcaagtagcg tatgcgctca cgcaactggt ccagaacctt gaccgaacgc agcggtggta
     5881 acggcgcagt ggcggttttc atggcttgtt atgactgttt ttttgtacag tctatgcctc
     5941 gggcatccaa gtagcaagcg cgttacgccg tgggtcgatg tttgatgtta tggagcagca
     6001 acgatgttac gcagcagggc agtcgcccta aaacaaagtt aggcagccgt tgtgctggtg
     6061 ctttctgata gttgttgtgg ggtaggcagt cagagctcga tttgcttgtc gccataatag
     6121 attcacaaga aggattcgac atgggtcaaa gtagcgatga agccaacgct cccgttgcag
     6181 ggcagtttgc gcttcccctg agtgccacct ttggcttagg ggatcgcgta cgcaagaaat
     6241 ctggtgccgc ttggcagggt caagtcgtcg gttggtattg cacaaaactc actcctgaag
     6301 gctatgcggt cgagtccgaa tcccacccag gctcagtgca aatttatcct gtggctgcac
     6361 ttgaacgtgt ggcctaagcg atatcttagg atctcccatc ggtgatgtcg gcgatatagg
     6421 cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga
     6481 tctgctcatg tttgacagct tatcatcgat gcataatgtg cctgtcaaat ggacgaagca
     6541 gggattctgc aaaccctatg ctactccgtc aagccgtcaa ttgtctgatt cgttaccaat
     6601 tatgacaact tgacggctac atcattcact ttttcttcac aaccggcacg gaactcgctc
     6661 gggctggccc cggtgcattt tttaaatacc cgcgagaaat agagttgatc gtcaaaacca
     6721 acattgcgac cgacggtggc gataggcatc cgggtggtgc tcaaaagcag cttcgcctgg
     6781 ctgatacgtt ggtcctcgcg ccagcttaag acgctaatcc ctaactgctg gcggaaaaga
     6841 tgtgacagac gcgacggcga caagcaaaca tgctgtgcga cgctggcgat atcaaaattg
     6901 ctgtctgcca ggtgatcgct gatgtactga caagcctcgc gtacccgatt atccatcggt
     6961 ggatggagcg actcgttaat cgcttccatg cgccgcagta acaattgctc aagcagattt
     7021 atcgccagca gctccgaata gcgcccttcc ccttgcccgg cgttaatgat ttgcccaaac
     7081 aggtcgctga aatgcggctg gtgcgcttca tccgggcgaa agaaccccgt attggcaaat
     7141 attgacggcc agttaagcca ttcatgccag taggcgcgcg gacgaaagta aacccactgg
     7201 tgataccatt cgcgagcctc cggatgacga ccgtagtgat gaatctctcc tggcgggaac
     7261 agcaaaatat cacccggtcg gcaaacaaat tctcgtccct gatttttcac caccccctga
     7321 ccgcgaatgg tgagattgag aatataacct ttcattccca gcggtcggtc gataaaaaaa
     7381 tcgagataac cgttggcctc aatcggcgtt aaacccgcca ccagatgggc attaaacgag
     7441 tatcccggca gcaggggatc attttgcgct tcagccatac ttttcatact cccgccattc
     7501 agag
//
