LOCUS       sequence-211988-        5516 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of ECFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5516)
  AUTHORS   Stuurman N, Pacios Bras C, Schlaman HR, Wijfjes AH, Bloemberg G, 
            Spaink HP
  TITLE     Use of green fluorescent protein color variants expressed on stable 
            broad-host-range vectors to visualize rhizobia interacting with 
            plants.
  JOURNAL   Mol Plant Microbe Interact. 2000 Nov;13(11):1163-9. doi: 
            10.1094/MPMI.2000.13.11.1163.
  PUBMED    11059482
REFERENCE   2  (bases 1 to 5516)
  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"; doi: 
            "10.1094/MPMI.2000.13.11"; journalName: "Mol Plant Microbe 
            Interact"; date: "2000-11"; volume: "13"; issue: "11"; pages: 
            "1163-9"
COMMENT     Sequence Label: pMP4516
FEATURES             Location/Qualifiers
     source          1..5516
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    155..176
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(191..196,197..214,215..221)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    229..245
                     /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)."
     primer_bind     234..256
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     253..269
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     253..269
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     288..308
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        290..308
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     337..353
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     338..354
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      406..415
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(412..414,415..417,418..1131)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(457..478)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(718..737)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1065..1086
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1181..1200)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1182..1200)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1210..1227)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1210..1226)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1219..1241)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(1935..2597)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      2598..3369
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             complement(4651..5184)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(5373..5378,5379..5395,5396..5401))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
ORIGIN
        1 gcttccatgt cggcagaatg cttaatgaat tacaacagtt tttatgcatg cgcccaatac
       61 gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc
      121 ccgactggaa agcgggcagt gagcgcaacg caattaatgt gagttagctc actcattagg
      181 caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat
      241 aacaatttca cacaggaaac agctatgacc atgattacgc caagcgcgca attaaccctc
      301 actaaaggga acaaaagctg ggtaccgggc cccccctcga ggtcgacggt atcgataagc
      361 ttgatatcga attcctgcag cccgggggat ccccgggtac cggtcgccac catggtgagc
      421 aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta
      481 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg
      541 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
      601 accctgacct ggggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac
      661 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac
      721 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
      781 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
      841 tacaactaca tcagccacaa cgtctatatc accgccgaca agcagaagaa cggcatcaag
      901 gccaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac
      961 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
     1021 acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
     1081 ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta aagcggccgc
     1141 gactctagag cggccgccac cgcggtggag ctccaattcg ccctatagtg agtcgtatta
     1201 cgcgcgctca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca
     1261 acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg
     1321 caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggaaat tgtaagcgtt
     1381 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     1441 gccgtactgc gatgagtggc agggcggggc gtaatttttt taaggcagtt attggtgccc
     1501 ttaaacgcct ggttgctacg cctgaataag tgataataag cggatgaatg gcagaaattc
     1561 gaaagcaaat tcgacccggt cgtcggttca gggcagggtc gttaaatagc cgcttatgtc
     1621 tattgctggt ttaccggttt attgactacc ggaagcagtg tgaccgtgtg cttctcaaat
     1681 gcctgaggcc agtttgctca ggctctcccc gtggaggtaa taattgacga tatgatcatt
     1741 tattctgcct cccagagcct gataaaaacg gtgaatccgt tagcgaggtg ccgccggctt
     1801 ccattcaggt cgaggtggcc cggctccatg caccgcgacg caacgcgggg aggcagacaa
     1861 ggtatagggc ggcgaggcgg ctacagccga tagtctggaa cagcgcactt acgggttgct
     1921 gcgcaaccca agtgctaccg gcgcggcagc gtgacccgtg tcggcggctc caacggctcg
     1981 ccatcgtcca gaaaacacgg ctcatcgggc atcggcaggc gctgctgccc gcgccgttcc
     2041 cattcctccg tttcggtcaa ggctggcagg tctggttcca tgcccggaat gccgggctgg
     2101 ctgggcggct cctcgccggg gccggtcggt agttgctgct cgcccggata cagggtcggg
     2161 atgcggcgca ggtcgccatg ccccaacagc gattcgtcct ggtcgtcgtg atcaaccacc
     2221 acggcggcac tgaacaccga caggcgcaac tggtcgcggg gctggcccca cgccacgcgg
     2281 tcattgacca cgtaggccga cacggtgccg gggccgttga gcttcacgac ggagatccag
     2341 cgctcggcca ccaagtcctt gactgcgtat tggaccgtcc gcaaagaacg tccgatgagc
     2401 ttggaaagtg tcttctggct gaccaccacg gcgttctggt ggcccatctg cgccacgagg
     2461 tgatgcagca gcattgccgc cgtgggtttc ctcgcaataa gcccggccca cgcctcatgc
     2521 gctttgcgtt ccgtttgcac ccagtgaccg ggcttgttct tggcttgaat gccgatttct
     2581 ctggactgcg tggccatgct tatctccatg cggtaggggt gccgcacggt tgcggcacca
     2641 tgcgcaatca gctgcaactt ttcggcagcg cgacaacaat tatgcgttgc gtaaaagtgg
     2701 cagtcaatta cagattttct ttaacctacg caatgagcta ttgcgggggg tgccgcaatg
     2761 agctgttgcg tacccccctt ttttaagttg ttgattttta agtctttcgc atttcgccct
     2821 atatctagtt ctttggtgcc caaagaaggg cacccctgcg gggttccccc acgccttcgg
     2881 cgcggctccc cctccggcaa aaagtggccc ctccggggct tgttgatcga ctgcgcggcc
     2941 ttcggccttg cccaaggtgg cgctgccccc ttggaacccc cgcactcgcc gccgtgaggc
     3001 tcggggggca ggcgggcggg cttcgccctt cgactgcccc cactcgcata ggcttgggtc
     3061 gttccaggcg cgtcaaggcc aagccgctgc gcggtcgctg cgcgagcctt gacccgcctt
     3121 ccacttggtg tccaaccggc aagcgaagcg cgcaggccgc aggccggagg cttttcccca
     3181 gagaaaatta aaaaaattga tggggcaagg ccgcaggccg cgcagttgga gccggtgggt
     3241 atgtggtcga aggctgggta gccggtgggc aatccctgtg gtcaagctcg tgggcaggcg
     3301 cagcctgtcc atcagcttgt ccagcagggt tgtccacggg ccgagcgaag cgagccagcc
     3361 ggtggccgct cgcggccatc gtccacatat ccacgggctg gcaagggagc gcagcgaccg
     3421 cgcagggcga agcccggaga gcaagcccgt agggcgccgc agccgccgta ggcggtcacg
     3481 actttgcgaa gcaaagtcta gtgagtatac tcaagcattg agtggcccgc cggaggcacc
     3541 gccttgcgct gcccccgtcg agccggttgg acaccaaaag ggaggggcag gcatggcggc
     3601 atacgcgatc atgcgatgca agaagctggc gaaaatgggc aacgtggcgg ccagtctcaa
     3661 gcacgcctac cgcgagcgcg agacgcccaa cgctgacgcc agcaggacgc cagagaacga
     3721 gcactgggcg gccagcagca ccgatgaagc gatgggccga ctgcgcgagt tgctgccaga
     3781 gaagcggcgc aaggacgctg tgttggcggt cgagtacgtc atgacggcca gcccggaatg
     3841 gtggaagtcg gccagccaag aacagcaggc ggcgttcttc gagaaggcgc acaagtggct
     3901 ggcggacaag tacggggcgg atcgcatcgt gacggccagc atccaccgtg acgaaaccag
     3961 cccgcacatg accgcgttcg tggtgccgct gacgcaggac ggcaggctgt cggccaagga
     4021 gttcatcggc aacaaagcgc agatgacccg cgaccagacc acgtttgcgg ccgctgtggc
     4081 cgatctaggg ctgcaacggg gcatcgaggg cagcaaggca cgtcacacgc gcattcaggc
     4141 gttctacgag gccctggagc ggccaccagt gggccacgtc accatcagcc cgcaagcggt
     4201 cgagccacgc gcctatgcac cgcagggatt ggccgaaaag ctgggaatct caaagcgcgt
     4261 tgagacgccg gaagccgtgg ccgaccggct gacaaaagcg gttcggcagg ggtatgagcc
     4321 tgccctacag gccgccgcag gagcgcgtga gatgcgcaag aaggccgatc aagcccaaga
     4381 gacggcccga gaccttcggg agcgcctgaa gcccgttctg gacgccctgg ggccgttgaa
     4441 tcgggatatg caggccaagg ccgccgcgat catcaaggcc gtgggcgaaa agctgctgac
     4501 ggaacagcgg gaagtccagc gccagaaaca ggcccagcgc cagcaggaac gcgggcgcgc
     4561 acatttcccc gaaaagtgcc acctgcggcg ttgtgacaat ttaccgaaca actccgcggc
     4621 cgggaagccg atctcggctt gaacgaattg ttaggtggcg gtacttgggt cgatatcaaa
     4681 gtgcatcact tcttcccgta tgcccaactt tgtatagaga gccactgcgg gatcgtcacc
     4741 gtaatctgct tgcacgtaga tcacataagc accaagcgcg ttggcctcat gcttgaggag
     4801 attgatgagc gcggtggcaa tgccctgcct ccggtgctct ccggagactg cgagatcata
     4861 gatatagatc tcactacgcg gctgctcaaa cttgggcaga acgtaagccg cgagagcgcc
     4921 aacaaccgct tcttggtcga aggcagcaag cgcgatgaat gtcttactac ggagcaagtt
     4981 cccgaggtaa tcggagtccg gctgatgttg ggagtaggtg gctacgtctc cgaactcacg
     5041 accgaaaaga tcaagagcag cccgcatgga tttgacttgg tcagggccga gcctacatgt
     5101 gcgaatgatg cccatacttg agccacctaa ctttgtttta gggcgactgc cctgctgcgt
     5161 aacatcgttg ctgctgcgta acatcgttgc tgctccataa catcaaacat cgacccacgg
     5221 cgtaacgcgc ttgctgcttg gatgcccgag gcatagactg tacaaaaaaa cagtcataac
     5281 aagccatgaa aaccgccact gcgccgttac caccgctgcg ttcggtcaag gttctggacc
     5341 agttgcgtga gcgcatacgc tacttgcatt acagtttacg aaccgaacag gcttatgtca
     5401 actgggttcg tgccttcatc cgtttccacg gtgtgcgtcc atgggcaaat attatacgca
     5461 aggcgacaag gtgctgatgc cgctggcgat tcaggttcat catgccgttt gtgatg
//
