pFC20K HaloTag T7 SP6 vector (V012500)

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LOCUS       40924_19826        4417 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Flexi(R) vector with a kanamycin resistance marker, for cell-free 
            expression of a protein with a cleavable C-terminal HaloTag(R).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4417)
  AUTHORS   Promega
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 4417)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Subclone a coding sequence between the SgfI/AsiSI and 
            EcoICRI/Eco53kI sites to remove the lethal barnase gene.
FEATURES             Location/Qualifiers
     source          1..4417
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        21..39
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        45..63
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     CDS             102..434
                     /codon_start=1
                     /label=barnase
                     /note="ribonuclease from Bacillus amyloliquefaciens"
                     /translation="MAQVINTFDGVADYLQTYHKLPDNYITKSEAQALGWVASKGNLAD
                     VAPGKSIGGDIFSNREGKLPGKSGRTWREADINYTSGFRNSDRILYSSDWLIYKTTDHY
                     QTFTKIR"
     CDS             477..497
                     /codon_start=1
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
                     /translation="EDLYFQS"
     CDS             510..1397
                     /codon_start=1
                     /label=HaloTag(R)
                     /note="modified bacterial dehalogenase that forms covalent
                     bonds with chloroalkane derivatives"
                     /translation="SEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSY
                     VWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIH
                     DWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIID
                     QNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVE
                     EYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLI
                     GSEIARWLSTLEISG"
     terminator      1540..1587
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             1968..2759
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase from Tn5"
                     /translation="MLEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     rep_origin      complement(2934..3522)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    3638..3921
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to
                     maintain plasmids as monomers"
     terminator      4151..4237
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      4329..4356
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"