LOCUS       sequence-206268-        4941 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pBBR1k-GFPuv with Anderson promoter J23101 expressing lacI.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4941)
  AUTHORS   Cook TB, Rand JM, Nurani W, Courtney DK, Liu SA, Pfleger BF
  TITLE     Genetic tools for reliable gene expression and recombineering in 
            Pseudomonas putida.
  JOURNAL   J Ind Microbiol Biotechnol. 2018 Jan 3. pii: 
            10.1007/s10295-017-2001-5. doi: 10.1007/s10295-017-2001-5.
  PUBMED    29299733
REFERENCE   2  (bases 1 to 4941)
  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: "J Ind 
            Microbiol Biotechnol. 2018 Jan 3. pii: 10.1007/s10295-017-2001-5. 
            doi: 10.1007/s10295-017-2001-5."
COMMENT     Sequence Label: pBBR1k-J23101-GFPuv
FEATURES             Location/Qualifiers
     source          1..4941
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      612..1383
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     terminator      complement(1417..1511)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(1542..2336)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1653..1672)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2263..2282
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        2491..2525
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     CDS             2552..3634
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(2571..2590)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(3866..3871,3872..3888,3889..3895)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    3903..3919
                     /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)."
     CDS             3960..4676
                     /codon_start=1
                     /product="GFP variant optimized for excitation by UV light"
                     /label=GFPuv
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(3990..4018)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     4613..4633
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      4709..4780
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4796..4823
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(join(4890..4941,1..611))
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
ORIGIN
        1 tccagaaaac acggctcatc gggcatcggc aggcgctgct gcccgcgccg ttcccattcc
       61 tccgtttcgg tcaaggctgg caggtctggt tccatgcccg gaatgccggg ctggctgggc
      121 ggctcctcgc cggggccggt cggtagttgc tgctcgcccg gatacagggt cgggatgcgg
      181 cgcaggtcgc catgccccaa cagcgattcg tcctggtcgt cgtgatcaac caccacggcg
      241 gcactgaaca ccgacaggcg caactggtcg cggggctggc cccacgccac gcggtcattg
      301 accacgtagg ccgacacggt gccggggccg ttgagcttca cgacggagat ccagcgctcg
      361 gccaccaagt ccttgactgc gtattggacc gtccgcaaag aacgtccgat gagcttggaa
      421 agtgtcttct ggctgaccac cacggcgttc tggtggccca tctgcgccac gaggtgatgc
      481 agcagcattg ccgccgtggg tttcctcgca ataagcccgg cccacgcctc atgcgctttg
      541 cgttccgttt gcacccagtg accgggcttg ttcttggctt gaatgccgat ttctctggac
      601 tgcgtggcca tgcttatctc catgcggtag gggtgccgca cggttgcggc accatgcgca
      661 atcagctgca acttttcggc agcgcgacaa caattatgcg ttgcgtaaaa gtggcagtca
      721 attacagatt ttctttaacc tacgcaatga gctattgcgg ggggtgccgc aatgagctgt
      781 tgcgtacccc ccttttttaa gttgttgatt tttaagtctt tcgcatttcg ccctatatct
      841 agttctttgg tgcccaaaga agggcacccc tgcggggttc ccccacgcct tcggcgcggc
      901 tccccctccg gcaaaaagtg gcccctccgg ggcttgttga tcgactgcgc ggccttcggc
      961 cttgcccaag gtggcgctgc ccccttggaa cccccgcact cgccgccgtg aggctcgggg
     1021 ggcaggcggg cgggcttcgc ccttcgactg cccccactcg cataggcttg ggtcgttcca
     1081 ggcgcgtcaa ggccaagccg ctgcgcggtc gctgcgcgag ccttgacccg ccttccactt
     1141 ggtgtccaac cggcaagcga agcgcgcagg ccgcaggccg gaggcttttc cccagagaaa
     1201 attaaaaaaa ttgatggggc aaggccgcag gccgcgcagt tggagccggt gggtatgtgg
     1261 tcgaaggctg ggtagccggt gggcaatccc tgtggtcaag ctcgtgggca ggcgcagcct
     1321 gtccatcagc ttgtccagca gggttgtcca cgggccgagc gaagcgagcc agccggtggc
     1381 cgctcgcggc catcgtccac atatcctagt gcttggattc tcaccaataa aaaacgcccg
     1441 gcggcaaccg agcgttctga acaaatccag atggagttct gaggtcatta ctggatctat
     1501 caacaggagt ccaagcgagc tctcgaaccc cagagtcccg ctcagaagaa ctcgtcaaga
     1561 aggcgataga aggcgatgcg ctgcgaatcg ggagcggcga taccgtaaag cacgaggaag
     1621 cggtcagccc attcgccgcc aagctcttca gcaatatcac gggtagccaa cgctatgtcc
     1681 tgatagcggt ccgccacacc cagccggcca cagtcgatga atccagaaaa gcggccattt
     1741 tccaccatga tattcggcaa gcaggcatcg ccatgggtca cgacgagatc ctcgccgtcg
     1801 ggcatgcgcg ccttgagcct ggcgaacagt tcggctggcg cgagcccctg atgctcttcg
     1861 tccagatcat cctgatcgac aagaccggct tccatccgag tacgtgctcg ctcgatgcga
     1921 tgtttcgctt ggtggtcgaa tgggcaggta gccggatcaa gcgtatgcag ccgccgcatt
     1981 gcatcagcca tgatggatac tttctcggca ggagcaaggt gagatgacag gagatcctgc
     2041 cccggcactt cgcccaatag cagccagtcc cttcccgctt cagtgacaac gtcgagcaca
     2101 gctgcgcaag gaacgcccgt cgtggccagc cacgatagcc gcgctgcctc gtcctgcagt
     2161 tcattcaggg caccggacag gtcggtcttg acaaaaagaa ccgggcgccc ctgcgctgac
     2221 agccggaaca cggcggcatc agagcagccg attgtctgtt gtgcccagtc atagccgaat
     2281 agcctctcca cccaagcggc cggagaacct gcgtgcaatc catcttgttc aatcatgcga
     2341 aacgatcctc atcctgtctc ttgatcagat catgatcccc tgcgccatca gatccttggc
     2401 ggcaagaaag ccatccagtt tactttgcag ggcttcccaa ccttaccaga gggcgcccca
     2461 gctggcaatt ccgacgtcga caccatcgaa tttacagcta gctcagtcct aggtattatg
     2521 ctagcaagag agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc
     2581 agagtatgcc ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt
     2641 ttctgcgaaa acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa
     2701 ccgcgtggca caacaactgg cgggcaaaca gtcgttgctg attggcgttg ccacctccag
     2761 tctggccctg cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact
     2821 gggtgccagc gtagtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc
     2881 ggtgcacaat cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga
     2941 ccaggatgcc attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt
     3001 ctctgaccag acacccatca acagtattat tttctcccat gaagacggta cgcgactggg
     3061 cgtggagcat ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag
     3121 ttctgtctcg gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat
     3181 tcagccgata gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat
     3241 gcaaatgctg aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc
     3301 gctgggcgca atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt
     3361 agtgggatac gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa
     3421 acaggatttt cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg
     3481 ccaggcggtg aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct
     3541 ggcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
     3601 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc
     3661 gcgaattgat ctggtttgac agcttatcat cgactgcacg gtgcaccaat gcttctggcg
     3721 tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg
     3781 tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt
     3841 ctggcaaata ttctgaaatg agctgttgac aattaatcat ccggctcgta taatgtgtgg
     3901 aattgtgagc ggataacaat ttcagaattc aaaagatctt ttaagaagga gatatacata
     3961 tgagtaaagg agaagaactt ttcactggag ttgtcccaat tcttgttgaa ttagatggtg
     4021 atgttaatgg gcacaaattt tctgtcagtg gagagggtga aggtgatgca acatacggaa
     4081 aacttaccct taaatttatt tgcactactg gaaaactacc tgttccgtgg ccaacacttg
     4141 tcactacttt ctcttatggt gttcaatgct tttcccgtta tccggatcac atgaaacggc
     4201 atgacttttt caagagtgcc atgcccgaag gttatgtaca ggaacgcact atatctttca
     4261 aagatgacgg gaactacaag acgcgtgctg aagtcaagtt tgaaggtgat acccttgtta
     4321 atcgtatcga gttaaaaggt attgatttta aagaagatgg aaacattctc ggacacaaac
     4381 tggagtacaa ctataactca cacaatgtat acatcacggc agacaaacaa aagaatggaa
     4441 tcaaagctaa cttcaaaatt cgccacaaca ttgaagatgg ctccgttcaa ctagcagacc
     4501 attatcaaca aaatactcca attggcgatg gccctgtcct tttaccagac aaccattacc
     4561 tgtccacaca atctgccctt tcgaaagatc ccaacgaaaa gcgtgaccac atggtccttc
     4621 ttgagtttgt aactgctgct gggattacac atggcatgga tgagctctac aaataaggat
     4681 ccaaactcga gtaaggatct ccaggcatca aataaaacga aaggctcagt cgaaagactg
     4741 ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc tactagagtc acactggctc
     4801 accttcgggt gggcctttct gcgtttatac ctaggctaca gccgatagtc tggaacagcg
     4861 cacttacggg ttgctgcgca acccaagtgc taccggcgcg gcagcgtgac ccgtgtcggc
     4921 ggctccaacg gctcgccatc g
//
