LOCUS       sequence-211994-        5516 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of EYFP.
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: pMP4518
FEATURES             Location/Qualifiers
     source          1..5516
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    245..266
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(281..286,287..304,305..311)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    319..335
                     /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     324..346
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     343..359
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     343..359
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     378..398
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        380..398
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     427..443
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     428..444
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      496..505
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(502..504,505..507,508..1221)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(547..568)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(808..827)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1155..1176
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1271..1290)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1272..1290)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1300..1317)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1300..1316)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1309..1331)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(2025..2687)
                     /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      2688..3459
                     /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(4741..5274)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(5463..5468,5469..5485,5486..5491))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
ORIGIN
        1 cacggtgtgc gtccatgggc aaatattata cgcaaggcga caaggtgctg atgccgctgg
       61 cgattcaggt tcatcatgcc gtttgtgatg gcttccatgt cggcagaatg cttaatgaat
      121 tacaacagtt tttatgcatg cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
      181 ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg
      241 caattaatgt gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg
      301 ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc
      361 atgattacgc caagcgcgca attaaccctc actaaaggga acaaaagctg ggtaccgggc
      421 cccccctcga ggtcgacggt atcgataagc ttgatatcga attcctgcag cccgggggat
      481 ccccgggtac cggtcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg
      541 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag
      601 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
      661 ctgcccgtgc cctggcccac cctcgtgacc accttcggct acggcgtgca gtgcttcgcc
      721 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
      781 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
      841 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
      901 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc
      961 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag
     1021 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     1081 gtgctgctgc ccgacaacca ctacctgagc taccagtccg ccctgagcaa agaccccaac
     1141 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     1201 atggacgagc tgtacaagta aagcggccgc gactctagag cggccgccac cgcggtggag
     1261 ctccaattcg ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg ttttacaacg
     1321 tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt
     1381 cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag
     1441 cctgaatggc gaatggaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt
     1501 tgttaaatca gctcattttt taaccaatag gccgtactgc gatgagtggc agggcggggc
     1561 gtaatttttt taaggcagtt attggtgccc ttaaacgcct ggttgctacg cctgaataag
     1621 tgataataag cggatgaatg gcagaaattc gaaagcaaat tcgacccggt cgtcggttca
     1681 gggcagggtc gttaaatagc cgcttatgtc tattgctggt ttaccggttt attgactacc
     1741 ggaagcagtg tgaccgtgtg cttctcaaat gcctgaggcc agtttgctca ggctctcccc
     1801 gtggaggtaa taattgacga tatgatcatt tattctgcct cccagagcct gataaaaacg
     1861 gtgaatccgt tagcgaggtg ccgccggctt ccattcaggt cgaggtggcc cggctccatg
     1921 caccgcgacg caacgcgggg aggcagacaa ggtatagggc ggcgaggcgg ctacagccga
     1981 tagtctggaa cagcgcactt acgggttgct gcgcaaccca agtgctaccg gcgcggcagc
     2041 gtgacccgtg tcggcggctc caacggctcg ccatcgtcca gaaaacacgg ctcatcgggc
     2101 atcggcaggc gctgctgccc gcgccgttcc cattcctccg tttcggtcaa ggctggcagg
     2161 tctggttcca tgcccggaat gccgggctgg ctgggcggct cctcgccggg gccggtcggt
     2221 agttgctgct cgcccggata cagggtcggg atgcggcgca ggtcgccatg ccccaacagc
     2281 gattcgtcct ggtcgtcgtg atcaaccacc acggcggcac tgaacaccga caggcgcaac
     2341 tggtcgcggg gctggcccca cgccacgcgg tcattgacca cgtaggccga cacggtgccg
     2401 gggccgttga gcttcacgac ggagatccag cgctcggcca ccaagtcctt gactgcgtat
     2461 tggaccgtcc gcaaagaacg tccgatgagc ttggaaagtg tcttctggct gaccaccacg
     2521 gcgttctggt ggcccatctg cgccacgagg tgatgcagca gcattgccgc cgtgggtttc
     2581 ctcgcaataa gcccggccca cgcctcatgc gctttgcgtt ccgtttgcac ccagtgaccg
     2641 ggcttgttct tggcttgaat gccgatttct ctggactgcg tggccatgct tatctccatg
     2701 cggtaggggt gccgcacggt tgcggcacca tgcgcaatca gctgcaactt ttcggcagcg
     2761 cgacaacaat tatgcgttgc gtaaaagtgg cagtcaatta cagattttct ttaacctacg
     2821 caatgagcta ttgcgggggg tgccgcaatg agctgttgcg tacccccctt ttttaagttg
     2881 ttgattttta agtctttcgc atttcgccct atatctagtt ctttggtgcc caaagaaggg
     2941 cacccctgcg gggttccccc acgccttcgg cgcggctccc cctccggcaa aaagtggccc
     3001 ctccggggct tgttgatcga ctgcgcggcc ttcggccttg cccaaggtgg cgctgccccc
     3061 ttggaacccc cgcactcgcc gccgtgaggc tcggggggca ggcgggcggg cttcgccctt
     3121 cgactgcccc cactcgcata ggcttgggtc gttccaggcg cgtcaaggcc aagccgctgc
     3181 gcggtcgctg cgcgagcctt gacccgcctt ccacttggtg tccaaccggc aagcgaagcg
     3241 cgcaggccgc aggccggagg cttttcccca gagaaaatta aaaaaattga tggggcaagg
     3301 ccgcaggccg cgcagttgga gccggtgggt atgtggtcga aggctgggta gccggtgggc
     3361 aatccctgtg gtcaagctcg tgggcaggcg cagcctgtcc atcagcttgt ccagcagggt
     3421 tgtccacggg ccgagcgaag cgagccagcc ggtggccgct cgcggccatc gtccacatat
     3481 ccacgggctg gcaagggagc gcagcgaccg cgcagggcga agcccggaga gcaagcccgt
     3541 agggcgccgc agccgccgta ggcggtcacg actttgcgaa gcaaagtcta gtgagtatac
     3601 tcaagcattg agtggcccgc cggaggcacc gccttgcgct gcccccgtcg agccggttgg
     3661 acaccaaaag ggaggggcag gcatggcggc atacgcgatc atgcgatgca agaagctggc
     3721 gaaaatgggc aacgtggcgg ccagtctcaa gcacgcctac cgcgagcgcg agacgcccaa
     3781 cgctgacgcc agcaggacgc cagagaacga gcactgggcg gccagcagca ccgatgaagc
     3841 gatgggccga ctgcgcgagt tgctgccaga gaagcggcgc aaggacgctg tgttggcggt
     3901 cgagtacgtc atgacggcca gcccggaatg gtggaagtcg gccagccaag aacagcaggc
     3961 ggcgttcttc gagaaggcgc acaagtggct ggcggacaag tacggggcgg atcgcatcgt
     4021 gacggccagc atccaccgtg acgaaaccag cccgcacatg accgcgttcg tggtgccgct
     4081 gacgcaggac ggcaggctgt cggccaagga gttcatcggc aacaaagcgc agatgacccg
     4141 cgaccagacc acgtttgcgg ccgctgtggc cgatctaggg ctgcaacggg gcatcgaggg
     4201 cagcaaggca cgtcacacgc gcattcaggc gttctacgag gccctggagc ggccaccagt
     4261 gggccacgtc accatcagcc cgcaagcggt cgagccacgc gcctatgcac cgcagggatt
     4321 ggccgaaaag ctgggaatct caaagcgcgt tgagacgccg gaagccgtgg ccgaccggct
     4381 gacaaaagcg gttcggcagg ggtatgagcc tgccctacag gccgccgcag gagcgcgtga
     4441 gatgcgcaag aaggccgatc aagcccaaga gacggcccga gaccttcggg agcgcctgaa
     4501 gcccgttctg gacgccctgg ggccgttgaa tcgggatatg caggccaagg ccgccgcgat
     4561 catcaaggcc gtgggcgaaa agctgctgac ggaacagcgg gaagtccagc gccagaaaca
     4621 ggcccagcgc cagcaggaac gcgggcgcgc acatttcccc gaaaagtgcc acctgcggcg
     4681 ttgtgacaat ttaccgaaca actccgcggc cgggaagccg atctcggctt gaacgaattg
     4741 ttaggtggcg gtacttgggt cgatatcaaa gtgcatcact tcttcccgta tgcccaactt
     4801 tgtatagaga gccactgcgg gatcgtcacc gtaatctgct tgcacgtaga tcacataagc
     4861 accaagcgcg ttggcctcat gcttgaggag attgatgagc gcggtggcaa tgccctgcct
     4921 ccggtgctct ccggagactg cgagatcata gatatagatc tcactacgcg gctgctcaaa
     4981 cttgggcaga acgtaagccg cgagagcgcc aacaaccgct tcttggtcga aggcagcaag
     5041 cgcgatgaat gtcttactac ggagcaagtt cccgaggtaa tcggagtccg gctgatgttg
     5101 ggagtaggtg gctacgtctc cgaactcacg accgaaaaga tcaagagcag cccgcatgga
     5161 tttgacttgg tcagggccga gcctacatgt gcgaatgatg cccatacttg agccacctaa
     5221 ctttgtttta gggcgactgc cctgctgcgt aacatcgttg ctgctgcgta acatcgttgc
     5281 tgctccataa catcaaacat cgacccacgg cgtaacgcgc ttgctgcttg gatgcccgag
     5341 gcatagactg tacaaaaaaa cagtcataac aagccatgaa aaccgccact gcgccgttac
     5401 caccgctgcg ttcggtcaag gttctggacc agttgcgtga gcgcatacgc tacttgcatt
     5461 acagtttacg aaccgaacag gcttatgtca actgggttcg tgccttcatc cgtttc
//
