LOCUS       sequence-122946-        7382 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 7382)
  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 7382)
  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[LVA]
FEATURES             Location/Qualifiers
     source          1..7382
                     /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      4844..4938
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             4982..5641
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(5048..5067)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      5706..5792
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(6131..6925)
                     /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(6242..6261)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     6852..6871
                     /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 ctatacaaaa ggcctgcagc aaacgacgaa
     4801 aactacgctt tagtagctta ataagcttaa ttagctgagc ttggactcct gttgatagat
     4861 ccagtaatga cctcagaact ccatctggat ttgttcagaa cgctcggttg ccgccgggcg
     4921 ttttttattg gtgagaatcc aagctagctt ggcgagattt tcaggagcta aggaagctaa
     4981 aatggagaaa aaaatcactg gatataccac cgttgatata tcccaatggc atcgtaaaga
     5041 acattttgag gcatttcagt cagttgctca atgtacctat aaccagaccg ttcagctgga
     5101 tattacggcc tttttaaaga ccgtaaagaa aaataagcac aagttttatc cggcctttat
     5161 tcacattctt gcccgcctga tgaatgctca tccggaattt cgtatggcaa tgaaagacgg
     5221 tgagctggtg atatgggata gtgttcaccc ttgttacacc gttttccatg agcaaactga
     5281 aacgttttca tcgctctgga gtgaatacca cgacgatttc cggcagtttc tacacatata
     5341 ttcgcaagat gtggcgtgtt acggtgaaaa cctggcctat ttccctaaag ggtttattga
     5401 gaatatgttt ttcgtctcag ccaatccctg ggtgagtttc accagttttg atttaaacgt
     5461 ggccaatatg gacaacttct tcgcccccgt tttcaccatg ggcaaatatt atacgcaagg
     5521 cgacaaggtg ctgatgccgc tggcgattca ggttcatcat gccgtctgtg atggcttcca
     5581 tgtcggcaga atgcttaatg aattacaaca gtactgcgat gagtggcagg gcggggcgta
     5641 atttttttaa ggcagttatt ggtgccctta aacgcctggg gtaatgactc tctagcttga
     5701 ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt
     5761 tgtcggtgaa cgctctcctg agtaggacaa atccgccgct aggagcttgc ggccgccagt
     5821 gtgatggata tctgcataaa aactgttgta attcattaag cattctgccg acatggaagc
     5881 catcacaaac ggcatgatga acctgaatcg ccagcggcat cagcaccttg tcgccttgcg
     5941 tataatattt gcccatgggg gtgggcgaag aactccagca tgagatcccc gcgctggagg
     6001 atcatccagc cggcgtcccg gaaaacgatt ccgaagccca acctttcata gaaggcggcg
     6061 gtggaatcga aatctcgtga tggcaggttg ggcgtcgctt ggtcggtcat ttcgaacccc
     6121 agagtcccgc tcagaagaac tcgtcaagaa ggcgatagaa ggcgatgcgc tgcgaatcgg
     6181 gagcggcgat accgtaaagc acgaggaagc ggtcagccca ttcgccgcca agctcttcag
     6241 caatatcacg ggtagccaac gctatgtcct gatagcggtc cgccacaccc agccggccac
     6301 agtcgatgaa tccagaaaag cggccatttt ccaccatgat attcggcaag caggcatcgc
     6361 catgggtcac gacgagatcc tcgccgtcgg gcatgcgcgc cttgagcctg gcgaacagtt
     6421 cggctggcgc gagcccctga tgctcttcgt ccagatcatc ctgatcgaca agaccggctt
     6481 ccatccgagt acgtgctcgc tcgatgcgat gtttcgcttg gtggtcgaat gggcaggtag
     6541 ccggatcaag cgtatgcagc cgccgcattg catcagccat gatggatact ttctcggcag
     6601 gagcaaggtg agatgacagg agatcctgcc ccggcacttc gcccaatagc agccagtccc
     6661 ttcccgcttc agtgacaacg tcgagcacag ctgcgcaagg aacgcccgtc gtggccagcc
     6721 acgatagccg cgctgcctcg tcctgcagtt cattcagggc accggacagg tcggtcttga
     6781 caaaaagaac cgggcgcccc tgcgctgaca gccggaacac ggcggcatca gagcagccga
     6841 ttgtctgttg tgcccagtca tagccgaata gcctctccac ccaagcggcc ggagaacctg
     6901 cgtgcaatcc atcttgttca atcatgcgaa acgatcctca tcctgtctct tgatcagatc
     6961 ttgatcccct gcgccatcag atccttggcg gcaagaaagc catccagttt actttgcagg
     7021 gcttcccaac cttaccagag ggcgccccag ctggcaattc cggttcgctt gctgtccata
     7081 aaaccgccca gtctagctat cgccatgtaa gcccactgca agctacctgc tttctctttg
     7141 cgcttgcgtt ttcccttgtc cagatagccc agtagctgac attcatccca ggtggcactt
     7201 ttcggggaaa tgtgcgcgcc cgcgttcctg ctggcgctgg gcctgtttct ggcgctggac
     7261 ttcccgctgt tccgtcagca gcttttcgcc cacggccttg atgatcgcgg cggccttggc
     7321 ctgcatatcc cgattcaacg gccccagggc gtccagaacg ggcttcaggc gctcccgaag
     7381 gt
//
