LOCUS       sequence-122945-        7340 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7340)
  AUTHORS   Miller WG, Leveau JH, Lindow SE
  TITLE     Improved gfp and inaZ broad-host-range promoter-probe vectors.
  JOURNAL   Mol Plant Microbe Interact. 2000 Nov;13(11):1243-50.
  PUBMED    11059491
REFERENCE   2  (bases 1 to 7340)
  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: "Mol Plant 
            Microbe Interact."; date: "2000-11"; volume: "13(11)"; pages: 
            "1243-50"
COMMENT     Sequence Label: pPROBE-gfp[tagless]
FEATURES             Location/Qualifiers
     source          1..7340
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1023..1792
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             1793..2455
                     /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     3147..3169
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3161..3178
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3162..3178
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3188..3207
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3188..3206
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      3247..3333
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3428..3514
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3609..3695
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3790..3876
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     misc_feature    3968..4024
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     CDS             4066..4779
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(4096..4124)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     4719..4739
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      4802..4896
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             4940..5599
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(5006..5025)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      5664..5750
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(6089..6883)
                     /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(6200..6219)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     6810..6829
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 ctcgggccgt ctcttgggct tgatcggcct tcttgcgcat ctcacgcgct cctgcggcgg
       61 cctgtagggc aggctcatac ccctgccgaa ccgcttttgt cagccggtcg gccacggctt
      121 ccggcgtctc aacgcgcttt gagattccca gcttttcggc caatccctgc ggtgcatagg
      181 cgcgtggctc gaccgcttgc gggctgatgg tgacgtggcc cactggtggc cgctccaggg
      241 cctcgtagaa cgcctgaatg cgcgtgtgac gtgccttgct gccctcgatg ccccgttgca
      301 gccctagatc ggccacagcg gccgcaaacg tggtctggtc gcgggtcatc tgcgctttgt
      361 tgccgatgaa ctccttggcc gacagcctgc cgtcctgcgt cagcggcacc acgaacgcgg
      421 tcatgtgcgg gctggtttcg tcacggtgga tgctggccgt cacgatgcga tccgccccgt
      481 acttgtccgc cagccacttg tgcgccttct cgaagaacgc cgcctgctgt tcttggctgg
      541 ccgacttcca ccattccggg ctggccgtca tgacgtactc gaccgccaac acagcgtcct
      601 tgcgccgctt ctctggcagc aactcgcgca gtcggcccat cgcttcatcg gtgctgctgg
      661 ccgcccagtg ctcgttctct ggcgtcctgc tggcgtcagc gttgggcgtc tcgcgctcgc
      721 ggtaggcgtg cttgagactg gccgccacgt tgcccatttt cgccagcttc ttgcatcgca
      781 tgatcgcgta tgccgccatg cctgcccctc ccttttggtg tccaaccggc tcgacggggg
      841 cagcgcaagg cggtgcctcc ggcgggccac tcaatgcttg agtatactca ctagactttg
      901 cttcgcaaag tcgtgaccgc ctacggcggc tgcggcgccc tacgggcttg ctctccgggc
      961 ttcgccctgc gcggtcgctg cgctcccttg ccagcccgtg gatatgtgga cgatggccgc
     1021 gagcggccac cggctggctc gcttcgctcg gcccgtggac aaccctgctg gacaagctga
     1081 tggacaggct gcgcctgccc acgagcttga ccacagggat tgcccaccgg ctacccagcc
     1141 ttcgaccaca tacccaccgg ctccaactgc gcggcctgcg gccttgcccc atcaattttt
     1201 ttaattttct ctggggaaaa gcctccggcc tgcggcctgc gcgcttcgct tgccggttgg
     1261 acaccaagtg gaaggcgggt caaggctcgc gcagcgaccg cgcagcggct tggccttgac
     1321 gcgcctggaa cgacccaagc ctatgcgagt gggggcagtc gaaggcgaag cccgcccgcc
     1381 tgccccccga gcctcacggc ggcgagtgcg ggggttccaa gggggcagcg ccaccttggg
     1441 caaggccgaa ggccgcgcag tcgatcaaca agccccggag gggccacttt ttgccggagg
     1501 gggagccgcg ccgaaggcgt gggggaaccc cgcaggggtg cccttctttg ggcaccaaag
     1561 aactagatat agggcgaaat gcgaaagact taaaaatcaa caacttaaaa aaggggggta
     1621 cgcaacagct cattgcggca ccccccgcaa tagctcattg cgtaggttaa agaaaatctg
     1681 taattgactg ccacttttac gcaacgcata attgttgtcg cgctgccgaa aagttgcagc
     1741 tgattgcgca tggtgccgca accgtgcggc accctaccgc atggagataa gcatggccac
     1801 gcagtccaga gaaatcggca ttcaagccaa gaacaagccc ggtcactggg tgcaaacgga
     1861 acgcaaagcg catgaggcgt gggccgggct tattgcgagg aaacccacgg cggcaatgct
     1921 gctgcatcac ctcgtggcgc agatgggcca ccagaacgcc gtggtggtca gccagaagac
     1981 actttccaag ctcatcggac gttctttgcg gacggtccaa tacgcagtca aggacttggt
     2041 ggccgagcgc tggatctccg tcgtgaagct caacggcccc ggcaccgtgt cggcctacgt
     2101 ggtcaatgac cgcgtggcgt ggggccagcc ccgcgaccag ttgcgcctgt cggtgttcag
     2161 tgccgccgtg gtggttgatc acgacgacca ggacgaatcg ctgttggggc atggcgacct
     2221 gcgccgcatc ccgaccctgt atccgggcga gcagcaacta ccgaccggcc ccggcgagga
     2281 gccgcccagc cagcccggca ttccgggcat ggaaccagac ctgccagcct tgaccgaaac
     2341 ggaggaatgg gaacggcgcg ggcagcagcg cctgccgatg cccgatgagc cgtgttttct
     2401 ggacgatggc gagccgttgg agccgccgac acgggtcacg ctgccgcgcc ggtagcactt
     2461 gggttgcgca gcaacccgta agtgcgctgt tccagactat cggctgtagc cgcctcgccg
     2521 ccctatacct tgtctgcctc cccgcgttgc gtcgcggtgc atggagccgg gccacctcga
     2581 cctgaatgga agccggcggc acctcgctaa cggattcacc gtttttatca ggctctggga
     2641 ggcagaataa atgatcatat cgtcaattat tacctccacg gggagagcct gagcaaactg
     2701 gcctcaggca tttgagaagc acacggtcac actgcttccg gtagtcaata aaccggtaaa
     2761 ccagcaatag acataagcgg ctatttaacg accctgccct gaaccgacga ccgggtcgaa
     2821 tttgctttcg aatttctgcc attcatccgc ttattatcac ttattcaggc gtagcaccag
     2881 gcgtttaagg gcaccaataa ctgccttaaa aaaattacgc cccgccctgc cactcatcgc
     2941 agtcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     3001 aaatattaac gcttacaatt tccattcgcc attcaggctg cgcaactgtt gggaagggcg
     3061 atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg
     3121 attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga
     3181 gcgcgcgtaa tacgactcac tatagggcga attggagctg gggccgcaat tcccaattcc
     3241 aggcatcaaa taaaacgaaa ggctcagtcg aaagactggg cctttcgttt tatctgttgt
     3301 ttgtcggtga acgctctcct gagtaggaca aatccgccgg gagcggattt gaacgttgcg
     3361 aagcaacggc ccggagggtg gcgggcagga cgcccgccat aaactgccag gaattaattc
     3421 caggcatcaa ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt ttatctgttg
     3481 tttgtcggtg aacgctctcc tgagtaggac aaatccgccg ggagcggatt tgaacgttgc
     3541 gaagcaacgg cccggagggt ggcgggcagg acgcccgcca taaactgcca ggaattaatt
     3601 ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt
     3661 gtttgtcggt gaacgctctc ctgagtagga caaatccgcc gggagcggat ttgaacgttg
     3721 cgaagcaacg gcccggaggg tggcgggcag gacgcccgcc ataaactgcc aggaattaat
     3781 tccaggcatc aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt
     3841 tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc cgggagcgga tttgaacgtt
     3901 gcgaagcaac ggcccggagg gtggcgggca ggacgcccgc cataaactgc caggaattgg
     3961 ggatcggaag cttgcatgcc tgcaggtcga ctctagagga tccccgggta ccgagctcga
     4021 attccctaac taactaaaga ttaactttat aaggaggaaa aacatatgag taaaggagaa
     4081 gaacttttca ctggagttgt cccaattctt gttgaattag atggtgatgt taatgggcac
     4141 aaattttctg tcagtggaga gggtgaaggt gatgcaacat acggaaaact tacccttaaa
     4201 tttatttgca ctactggaaa actacctgtt ccatggccaa cacttgtcac tactttgact
     4261 tatggtgttc aatgcttttc aagataccca gatcatatga aacggcatga ctttttcaag
     4321 agtgccatgc ccgaaggtta tgtacaggaa agaactatat ttttcaaaga tgacgggaac
     4381 tacaagacac gtgctgaagt caagtttgaa ggtgataccc ttgttaatag aatcgagtta
     4441 aaaggtattg attttaaaga agatggaaac attcttggac acaaattgga atacaactat
     4501 aactcacaca atgtatacat catggcagac aaacaaaaga atggaatcaa agttaacttc
     4561 aaaattagac acaacattga agatggaagc gttcaactag cagaccatta tcaacaaaat
     4621 actccaattg gcgatggccc tgtcctttta ccagacaacc attacctgtc cacacaatct
     4681 gccctttcga aagatcccaa cgaaaagaga gaccacatgg tccttcttga gtttgtaaca
     4741 gctgctggga ttacacatgg catggatgaa ctatacaaat aagcttaatt agctgagctt
     4801 ggactcctgt tgatagatcc agtaatgacc tcagaactcc atctggattt gttcagaacg
     4861 ctcggttgcc gccgggcgtt ttttattggt gagaatccaa gctagcttgg cgagattttc
     4921 aggagctaag gaagctaaaa tggagaaaaa aatcactgga tataccaccg ttgatatatc
     4981 ccaatggcat cgtaaagaac attttgaggc atttcagtca gttgctcaat gtacctataa
     5041 ccagaccgtt cagctggata ttacggcctt tttaaagacc gtaaagaaaa ataagcacaa
     5101 gttttatccg gcctttattc acattcttgc ccgcctgatg aatgctcatc cggaatttcg
     5161 tatggcaatg aaagacggtg agctggtgat atgggatagt gttcaccctt gttacaccgt
     5221 tttccatgag caaactgaaa cgttttcatc gctctggagt gaataccacg acgatttccg
     5281 gcagtttcta cacatatatt cgcaagatgt ggcgtgttac ggtgaaaacc tggcctattt
     5341 ccctaaaggg tttattgaga atatgttttt cgtctcagcc aatccctggg tgagtttcac
     5401 cagttttgat ttaaacgtgg ccaatatgga caacttcttc gcccccgttt tcaccatggg
     5461 caaatattat acgcaaggcg acaaggtgct gatgccgctg gcgattcagg ttcatcatgc
     5521 cgtctgtgat ggcttccatg tcggcagaat gcttaatgaa ttacaacagt actgcgatga
     5581 gtggcagggc ggggcgtaat ttttttaagg cagttattgg tgcccttaaa cgcctggggt
     5641 aatgactctc tagcttgagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct
     5701 ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag taggacaaat ccgccgctag
     5761 gagcttgcgg ccgccagtgt gatggatatc tgcataaaaa ctgttgtaat tcattaagca
     5821 ttctgccgac atggaagcca tcacaaacgg catgatgaac ctgaatcgcc agcggcatca
     5881 gcaccttgtc gccttgcgta taatatttgc ccatgggggt gggcgaagaa ctccagcatg
     5941 agatccccgc gctggaggat catccagccg gcgtcccgga aaacgattcc gaagcccaac
     6001 ctttcataga aggcggcggt ggaatcgaaa tctcgtgatg gcaggttggg cgtcgcttgg
     6061 tcggtcattt cgaaccccag agtcccgctc agaagaactc gtcaagaagg cgatagaagg
     6121 cgatgcgctg cgaatcggga gcggcgatac cgtaaagcac gaggaagcgg tcagcccatt
     6181 cgccgccaag ctcttcagca atatcacggg tagccaacgc tatgtcctga tagcggtccg
     6241 ccacacccag ccggccacag tcgatgaatc cagaaaagcg gccattttcc accatgatat
     6301 tcggcaagca ggcatcgcca tgggtcacga cgagatcctc gccgtcgggc atgcgcgcct
     6361 tgagcctggc gaacagttcg gctggcgcga gcccctgatg ctcttcgtcc agatcatcct
     6421 gatcgacaag accggcttcc atccgagtac gtgctcgctc gatgcgatgt ttcgcttggt
     6481 ggtcgaatgg gcaggtagcc ggatcaagcg tatgcagccg ccgcattgca tcagccatga
     6541 tggatacttt ctcggcagga gcaaggtgag atgacaggag atcctgcccc ggcacttcgc
     6601 ccaatagcag ccagtccctt cccgcttcag tgacaacgtc gagcacagct gcgcaaggaa
     6661 cgcccgtcgt ggccagccac gatagccgcg ctgcctcgtc ctgcagttca ttcagggcac
     6721 cggacaggtc ggtcttgaca aaaagaaccg ggcgcccctg cgctgacagc cggaacacgg
     6781 cggcatcaga gcagccgatt gtctgttgtg cccagtcata gccgaatagc ctctccaccc
     6841 aagcggccgg agaacctgcg tgcaatccat cttgttcaat catgcgaaac gatcctcatc
     6901 ctgtctcttg atcagatctt gatcccctgc gccatcagat ccttggcggc aagaaagcca
     6961 tccagtttac tttgcagggc ttcccaacct taccagaggg cgccccagct ggcaattccg
     7021 gttcgcttgc tgtccataaa accgcccagt ctagctatcg ccatgtaagc ccactgcaag
     7081 ctacctgctt tctctttgcg cttgcgtttt cccttgtcca gatagcccag tagctgacat
     7141 tcatcccagg tggcactttt cggggaaatg tgcgcgcccg cgttcctgct ggcgctgggc
     7201 ctgtttctgg cgctggactt cccgctgttc cgtcagcagc ttttcgccca cggccttgat
     7261 gatcgcggcg gccttggcct gcatatcccg attcaacggc cccagggcgt ccagaacggg
     7321 cttcaggcgc tcccgaaggt
//
