LOCUS       sequence-193292-        7358 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Pseudomonas T3SS gateway vector with HA-sfGFP11-sfGFP11.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7358)
  AUTHORS   Park E, Lee HY, Woo J, Choi D, Dinesh-Kumar SP
  TITLE     Spatiotemporal Monitoring of Pseudomonas syringae Effectors via Type
            III Secretion Using Split Fluorescent Protein Fragments.
  JOURNAL   Plant Cell. 2017 Jul;29(7):1571-1584. doi: 10.1105/tpc.17.00047. 
            Epub 2017 Jun 14.
  PUBMED    28619883
REFERENCE   2  (bases 1 to 7358)
  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: 
            "10.1105/tpc.17.00047"; journalName: "Plant Cell"; date: "2017-07"; 
            volume: "29"; issue: "7"; pages: "1571-1584"
COMMENT     Sequence Label: PEP119T
FEATURES             Location/Qualifiers
     source          1..7358
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1934..2728
                     /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(1988..2007)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2598..2617
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     protein_bind    3133..3154
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3169..3174,3175..3192,3193..3199)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3207..3223
                     /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     3212..3234
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3231..3247
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3231..3247
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     3266..3286
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        3268..3286
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             complement(3501..3548)
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     CDS             complement(3594..3641)
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     CDS             complement(3687..3713)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    3715..3839
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(3880..4185)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(3982..4001)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(4527..5186)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     5101..5120
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(5240..5246,5247..5264,5265..5270))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(5295..5419)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(5624..5643)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5625..5643)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5653..5670)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5653..5669)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5662..5684)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(6378..7040)
                     /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      join(7041..7358,1..454)
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
ORIGIN
        1 gggcttgttg atcgactgcg cggccttcgg ccttgcccaa ggtggcgctg cccccttgga
       61 acccccgcac tcgccgccgt gaggctcggg gggcaggcgg gcgggcttcg cccttcgact
      121 gcccccactc gcataggctt gggtcgttcc aggcgcgtca aggccaagcc gctgcgcggt
      181 cgctgcgcga gccttgaccc gccttccact tggtgtccaa ccggcaagcg aagcgcgcag
      241 gccgcaggcc ggaggctttt ccccagagaa aattaaaaaa attgatgggg caaggccgca
      301 ggccgcgcag ttggagccgg tgggtatgtg gtcgaaggct gggtagccgg tgggcaatcc
      361 ctgtggtcaa gctcgtgggc aggcgcagcc tgtccatcag cttgtccagc agggttgtcc
      421 acgggccgag cgaagcgagc cagccggtgg ccgctcgcgg ccatcgtcca catatccacg
      481 ggctggcaag ggagcgcagc gaccgcgcag ggcgaagccc ggagagcaag cccgtagggc
      541 gccgcagccg ccgtaggcgg tcacgacttt gcgaagcaaa gtctagtgag tatactcaag
      601 cattgagtgg cccgccggag gcaccgcctt gcgctgcccc cgtcgagccg gttggacacc
      661 aaaagggagg ggcaggcatg gcggcatacg cgatcatgcg atgcaagaag ctggcgaaaa
      721 tgggcaacgt ggcggccagt ctcaagcacg cctaccgcga gcgcgagacg cccaacgctg
      781 acgccagcag gacgccagag aacgagcact gggcggccag cagcaccgat gaagcgatgg
      841 gccgactgcg cgagttgctg ccagagaagc ggcgcaagga cgctgtgttg gcggtcgagt
      901 acgtcatgac ggccagcccg gaatggtgga agtcggccag ccaagaacag caggcggcgt
      961 tcttcgagaa ggcgcacaag tggctggcgg acaagtacgg ggcggatcgc atcgtgacgg
     1021 ccagcatcca ccgtgacgaa accagcccgc acatgaccgc gttcgtggtg ccgctgacgc
     1081 aggacggcag gctgtcggcc aaggagttca tcggcaacaa agcgcagatg acccgcgacc
     1141 agaccacgtt tgcggccgct gtggccgatc tagggctgca acggggcatc gagggcagca
     1201 aggcacgtca cacgcgcatt caggcgttct acgaggccct ggagcggcca ccagtgggcc
     1261 acgtcaccat cagcccgcaa gcggtcgagc cacgcgccta tgcaccgcag ggattggccg
     1321 aaaagctggg aatctcaaag cgcgttgaga cgccggaagc cgtggccgac cggctgacaa
     1381 aagcggttcg gcaggggtat gagtctgccc tacaggccgc cgcaggagcg cgtgagatgc
     1441 gcaagaaggc cgatcaagcc caagagacgg cccgagacct tcgggagcgc ctgaagcccg
     1501 ttctggacgc cctggggccg ttgaatcggg atatgcaggc caaggccgcc gcgatcatca
     1561 aggccgtggg cgaaaagctg ctgacggaac agcgggaagt ccagcgccag aaacaggccc
     1621 agcgccagca ggaacgcggg cgcgcacatt tccccgaaaa gtgccacctg ggatgaatgt
     1681 cagctactgg gctatctgga caagggaaaa cgcaagcgca aagagaaagc aggtagcttg
     1741 cagtgggctt acatggcgat agctagactg ggcggtttta tggacagcaa gcgaaccgga
     1801 attgccagct ggggcgccct ctggtaaggt tgggaagccc tgcaaagtaa actggatggc
     1861 tttcttgccg ccaaggatct gatggcgcag gggatcaaga tctgatcaag agacaggatg
     1921 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     1981 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     2041 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     2101 cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     2161 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     2221 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     2281 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     2341 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     2401 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     2461 gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     2521 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     2581 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     2641 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     2701 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa
     2761 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
     2821 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     2881 ctggagttct tcgcccaccc ccatgggcaa atattatacg caaggcgaca aggtgctgat
     2941 gccgctggcg attcaggttc atcatgccgt ttgtgatggc ttccatgtcg gcagaatgct
     3001 taatgaatta caacagtttt tatgcatgcg cccaatacgc aaaccgcctc tccccgcgcg
     3061 ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga
     3121 gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat
     3181 gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag
     3241 ctatgaccat gattacgcca agcgcgcaat taaccctcac taaagggaac aaaagctggg
     3301 tacctcatgg gtattctgat ttggtgtttg tcctgaggaa ctctgaagaa gacgtccaaa
     3361 acagagcgac tgtcgatgcg aagccatagc taaagctgat atctatgttt catacctatt
     3421 ttcttcagtc accgttttga aaaccttacc tccctggcaa ggtcgtatta atgatttttg
     3481 gtagccttat cctcgagtta agtgatgcct gctgcgttta cgtattcgtg gagaaccata
     3541 tggtctcgcg aactgcctcc acccgagcct ccacctccac ttccgccgcc acctgtgatg
     3601 ccggcggcgt tcacgtactc gtgcagcacc atgtggtccc ggctccctcc gccaccgctc
     3661 cctccgccac cggatccgcc gccgccagcg tagtctggga cgtcgtatgg gtagaccact
     3721 ttgtacaaga aagctgaacg agaaacgtaa aatgatataa atatcaatat attaaattag
     3781 attttgcata aaaaacagac tacataatac tgtaaaacac aacatatcca gtcactatgg
     3841 tcgacctgca gactggctgt gtataaggga gcctgacatt tatattcccc agaacatcag
     3901 gttaatggcg tttttgatgt cattttcgcg gtggctgaga tcagccactt cttccccgat
     3961 aacggagacc ggcacactgg ccatatcggt ggtcatcatg cgccagcttt catccccgat
     4021 atgcaccacc gggtaaagtt cacgggagac tttatctgac agcagacgtg cactggccag
     4081 ggggatcacc atccgtcgcc cgggcgtgtc aataatatca ctctgtacat ccacaaacag
     4141 acgataacgg ctctctcttt tataggtgta aaccttaaac tgcatttcac cagcccctgt
     4201 tctcgtcagc aaaagagccg ttcatttcaa taaaccgggc gacctcagcc atcccttcct
     4261 gattttccgc tttccagcgt tcggcacgca gacgacgggc ttcattctgc atggttgtgc
     4321 ttaccagacc ggagatattg acatcatata tgccttgagc aactgatagc tgtcgctgtc
     4381 aactgtcact gtaatacgct gcttcataac atacctcttt ttgacatact tcgggtatac
     4441 atatcagtat atattcttat accgcaaaaa tcagcgcgca aatacgcata ctgttatctg
     4501 gcttttagta agccggatcc acgcgtttac gccccgccct gccactcatc gcagtactgt
     4561 tgtaattcat taagcattct gccgacatgg aagccatcac aaacggcatg atgaacctga
     4621 atcgccagcg gcatcagcac cttgtcgcct tgcgtataat atttgcccat ggtgaaaacg
     4681 ggggcgaaga agttgtccat attggccacg tttaaatcaa aactggtgaa actcacccag
     4741 ggattggctg agacgaaaaa catattctca ataaaccctt tagggaaata ggccaggttt
     4801 tcaccgtaac acgccacatc ttgcgaatat atgtgtagaa actgccggaa atcgtcgtgg
     4861 tattcactcc agagcgatga aaacgtttca gtttgctcat ggaaaacggt gtaacaaggg
     4921 tgaacactat cccatatcac cagctcaccg tctttcattg ccatacggaa ttccggatga
     4981 gcattcatca ggcgggcaag aatgtgaata aaggccggat aaaacttgtg cttatttttc
     5041 tttacggtct ttaaaaaggc cgtaatatcc agctgaacgg tctggttata ggtacattga
     5101 gcaactgact gaaatgcctc aaaatgttct ttacgatgcc attgggatat atcaacggtg
     5161 gtatatccag tgattttttt ctccatttta gcttccttag ctcctgaaaa tctcgacgga
     5221 tcctaactca aaatccacac attatacgag ccggaagcat aaagtgtaaa gcctggggtg
     5281 cctaatgcgg ccgccatagt gactggatat gttgtgtttt acagtattat gtagtctgtt
     5341 ttttatgcaa aatctaattt aatatattga tatttatatc attttacgtt tctcgttcag
     5401 cttttttgta caaacttgtc ctaggaaaaa accttttgat tgaaagtatg gagtacatgt
     5461 gtggttttaa aagaaaatga gttccctgct ggaacagact ccattgggcg gaaaacgatt
     5521 ttttagactg cgcatactcg cctgcgtcca tcacaaatct acagatctga acaggctcag
     5581 gccgaaaaag cctgcgtacg atttttgtct gagctccaat tcgccctata gtgagtcgta
     5641 ttacgcgcgc tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac
     5701 ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc
     5761 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatgga aattgtaagc
     5821 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     5881 taggccgtac tgcgatgagt ggcagggcgg ggcgtaattt ttttaaggca gttattggtg
     5941 cccttaaacg cctggttgct acgcctgaat aagtgataat aagcggatga atggcagaaa
     6001 ttcgaaagca aattcgaccc ggtcgtcggt tcagggcagg gtcgttaaat agccgcttat
     6061 gtctattgct ggtttaccgg tttattgact accggaagca gtgtgaccgt gtgcttctca
     6121 aatgcctgag gccagtttgc tcaggctctc cccgtggagg taataattga cgatatgatc
     6181 atttattctg cctcccagag cctgataaaa acggtgaatc cgttagcgag gtgccgccgg
     6241 cttccattca ggtcgaggtg gcccggctcc atgcaccgcg acgcaacgcg gggaggcaga
     6301 caaggtatag ggcggcgagg cggctacagc cgatagtctg gaacagcgca cttacgggtt
     6361 gctgcgcaac ccaagtgcta ccggcgcggc agcgtgaccc gtgtcggcgg ctccaacggc
     6421 tcgccatcgt ccagaaaaca cggctcatcg ggcatcggca ggcgctgctg cccgcgccgt
     6481 tcccattcct ccgtttcggt caaggctggc aggtctggtt ccatgcccgg aatgccgggc
     6541 tggctgggcg gctcctcgcc ggggccggtc ggtagttgct gctcgcccgg atacagggtc
     6601 gggatgcggc gcaggtcgcc atgccccaac agcgattcgt cctggtcgtc gtgatcaacc
     6661 accacggcgg cactgaacac cgacaggcgc aactggtcgc ggggctggcc ccacgccacg
     6721 cggtcattga ccacgtaggc cgacacggtg ccggggccgt tgagcttcac gacggagatc
     6781 cagcgctcgg ccaccaagtc cttgactgcg tattggaccg tccgcaaaga acgtccgatg
     6841 agcttggaaa gtgtcttctg gctgaccacc acggcgttct ggtggcccat ctgcgccacg
     6901 aggtgatgca gcagcattgc cgccgtgggt ttcctcgcaa taagcccggc ccacgcctca
     6961 tgcgctttgc gttccgtttg cacccagtga ccgggcttgt tcttggcttg aatgccgatt
     7021 tctctggact gcgtggccat gcttatctcc atgcggtagg ggtgccgcac ggttgcggca
     7081 ccatgcgcaa tcagctgcaa cttttcggca gcgcgacaac aattatgcgt tgcgtaaaag
     7141 tggcagtcaa ttacagattt tctttaacct acgcaatgag ctattgcggg gggtgccgca
     7201 atgagctgtt gcgtaccccc cttttttaag ttgttgattt ttaagtcttt cgcatttcgc
     7261 cctatatcta gttctttggt gcccaaagaa gggcacccct gcggggttcc cccacgcctt
     7321 cggcgcggct ccccctccgg caaaaagtgg cccctccg
//
