LOCUS       Exported                3684 bp ds-DNA    circular SYN 27-6-2015
DEFINITION  Flexi(R) vector with a kanamycin resistance marker, for cell-free 
            translation in a wheat germ extract.
ACCESSION   AY949044
VERSION     .
KEYWORDS    pF3K WG (BYDV)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3684)
  AUTHORS   Promega
  TITLE     Direct Submission
COMMENT     Subclone a coding sequence between the SgfI/AsiSI and PmeI sites to 
            remove the lethal barnase gene.
FEATURES             Location/Qualifiers
     source          1..3684
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        23..41
                     /note="SP6 promoter"
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     promoter        46..63
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     5UTR           65..201
                     /note="BYDV 5 UTR"
                     /note="barley yellow dwarf virus 5 untranslated region; 
                     required for function of the cap-independent 3 translation
                     enhancer (Guo et al., 2001)"
     CDS             233..568
                     /codon_start=1
                     /product="ribonuclease from Bacillus amyloliquefaciens"
                     /note="barnase"
                     /note="The barnase gene is lethal in standard bacterial 
                     transformation strains."
                     /protein_id="
                     "
                     /translation="MAQVINTFDGVADYLQTYHKLPDNYITKSEAQALGWVASKGNLAD
                     VAPGKSIGGDIFSNREGKLQGKSGRTWREADINYTSGFRNSDRILYSSDWLIYKTTDHY
                     QTFTKIR"
     misc_feature    596..702
                     /note="BYDV 3 TE"
                     /note="cap-independent 3 translation enhancer from barley 
                     yellow dwarf virus (Wang et al., 1997)"
     terminator      807..854
                     /note="T7 terminator"
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             1235..2029
                     /codon_start=1
                     /gene="aph(3)-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /note="NeoR/KanR"
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MLEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     rep_origin      complement(2201..2789)
                     /direction=LEFT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    2905..3188
                     /note="cer region"
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
     terminator      3418..3504
                     /gene="Escherichia coli rrnB"
                     /note="rrnB T1 terminator"
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3596..3623
                     /note="rrnB T2 terminator"
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"