LOCUS       Exported               18691 bp ds-DNA    circular SYN 01-12-2013
DEFINITION  High-throughput binary Gateway(R) donor vector for silencing genes 
            in plants using intron-containing hairpin RNA.
ACCESSION   AJ311874
VERSION     .
KEYWORDS    pHELLSGATE
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 18691)
  AUTHORS   Wesley SV, Helliwell CA, Smith NA, Wang MB, Rouse DT, Liu Q, Gooding
            PS, Singh SP, Abbott D, Stoutjesdijk PA, Robinson SP, Gleave AP, 
            Green AG, Waterhouse PM.
  TITLE     Construct design for efficient, effective and high-throughput gene 
            silencing in plants.
  JOURNAL   Plant J. 2001;27:581-90.
  PUBMED    11576441
REFERENCE   2  (bases 1 to 18691)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..18691
                     /organism="synthetic DNA construct"
                     /lab_host="Plant Cells"
                     /mol_type="other DNA"
     promoter        264..447
                     /function="NOS promoter"
                     /note="NOS promoter"
                     /note="nopaline synthase promoter"
     CDS             448..1269
                     /codon_start=1
                     /gene="nos-nptII"
                     /product="fusion between an N-terminal peptide of nopaline 
                     synthase and Tn5 aminoglycoside phosphotransferase"
                     /note="KanR"
                     /note="confers resistance to kanamycin"
                     /protein_id="
                     "
                     /translation="MAITLSATSLPISARIRAGSPAAWVERLFGYDWAQQTIGCSDAAV
                     FRLSAQGRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLG
                     EVPGQDLLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVD
                     QDDLDEEHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLG
                     VADRYQDIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     terminator      1894..2146
                     /note="NOS terminator"
                     /note="nopaline synthase terminator and poly(A) signal"
     misc_feature    2268..2292
                     /note="LB T-DNA repeat"
                     /note="left border repeat from nopaline C58 T-DNA"
     oriT            2846..2955
                     /note="incP origin of transfer"
     mobile_element  3015..3782
                     /mobile_element_type="insertion sequence:IS1"
                     /note="IS1"
                     /note="prokaryotic transposable element"
     CDS             4407..5555
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /note="trfA"
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     MVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     rep_origin      complement(6915..7503)
                     /direction=LEFT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             8600..9388
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase"
                     /note="SmR"
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      10071..10684
                     /note="oriV"
                     /note="origin of replication for the bacterial F plasmid"
     misc_feature    11182..11206
                     /note="RB T-DNA repeat"
                     /note="right border repeat from nopaline C58 T-DNA"
     promoter        12672..13017
                     /note="CaMV 35S promoter"
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     protein_bind    13048..13279
                     /gene="mutant version of attP"
                     /bound_moiety="BP Clonase(TM)"
                     /note="attP1"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(13675..13980)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /note="ccdB"
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(14387..14618)
                     /gene="mutant version of attP"
                     /bound_moiety="BP Clonase(TM)"
                     /note="attP2"
                     /note="recombination site for the Gateway(R) BP reaction 
                     (pDONR(TM)221 version)"
     intron          14662..16258
                     /gene="pdk"
                     /number="3"
                     /note="PDK intron"
                     /note="modified pyruvate orthophosphate dikinase intron 
                     from Flaveria trinervia"
     CDS             15002..15661
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /note="CmR"
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     protein_bind    16320..16551
                     /gene="mutant version of attP"
                     /bound_moiety="BP Clonase(TM)"
                     /note="attP2"
                     /note="recombination site for the Gateway(R) BP reaction 
                     (pDONR(TM)221 version)"
     CDS             complement(16855..17160)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /note="ccdB"
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(17659..17890)
                     /gene="mutant version of attP"
                     /bound_moiety="BP Clonase(TM)"
                     /note="attP1"
                     /note="recombination site for the Gateway(R) BP reaction"
     terminator      17922..18628
                     /note="OCS terminator"
                     /note="octopine synthase terminator"