pHELLSGATE 12 vector (V011791)

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LOCUS       pHELLSGATE_12.       17681 bp DNA     circular SYN 01-JAN-1980
DEFINITION  High-throughput binary Gateway(R) destination vector with two 
            introns, for silencing genes in plants using intron-containing 
            hairpin RNA.
ACCESSION   .
VERSION     .
KEYWORDS    pHELLSGATE 12
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 17681)
  AUTHORS   Helliwell CA, Waterhouse PM.
  TITLE     Constructs and methods for hairpin RNA-mediated gene silencing in 
            plants.
  JOURNAL   Meth. Enzymol. 2005;392:24-35.
  PUBMED    15644173
REFERENCE   2  (bases 1 to 17681)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 17681)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Meth. 
            Enzymol."; date: "2005"; volume: "392"; pages: "24-35"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..17681
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(65..83)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(90..106)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        264..447
                     /label=NOS promoter
                     /note="nopaline synthase promoter"
     CDS             448..1266
                     /label=KanR
                     /note="fusion between an N-terminal peptide of nopaline
                     synthase and Tn5 aminoglycoside phosphotransferase"
     terminator      1894..2146
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     misc_feature    complement(2268..2292)
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     oriT            2846..2955
                     /label=oriT
                     /note="incP origin of transfer"
     mobile_element  3015..3782
                     /label=IS1
                     /note="prokaryotic transposable element"
     CDS             3782..4132
                     /label=traJ
                     /note="oriT-recognizing protein"
     CDS             4407..5552
                     /label=trfA
                     /note="trans-acting replication protein that binds to and 
                     activates oriV"
     rep_origin      complement(6915..7503)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             8600..9388
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase"
                     /label=aadA
                     /note="SmR"
                     /note="confers resistance to spectinomycin and
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      complement(9993..10705)
                     /direction=LEFT
                     /label=oriV
                     /note="incP origin of replication"
     misc_feature    complement(11182..11206)
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     protein_bind    11462..11483
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        11498..11528
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11536..11552
                     /label=lac operator
                     /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     11560..11576
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        11594..11612
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     promoter        12672..13017
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     protein_bind    13026..13150
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(13582..13884)
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
     protein_bind    complement(14324..14448)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     intron          14475..15241
                     /label=PDK intron
                     /note="pyruvate orthophosphate dikinase intron from
                     Flaveria trinervia"
     intron          complement(15276..15465)
                     /label=cat1 intron
                     /note="castor bean catalase intron, modified"
     protein_bind    15489..15613
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(15945..16247)
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
     protein_bind    complement(16779..16903)
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     terminator      16912..17618
                     /label=OCS terminator
                     /note="octopine synthase terminator"
     promoter        complement(17659..17677)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"