This plasmid is designed to express tagged proteins in E. coli. The plasmid contains a constitutive promoter (OXB20) derived from the region upstream of the E. coli RecA gene. It does not require induction or any additional components for activity. It is the strongest of the bacterial promoters that we provide and this high level of expression can cause expression problems with some proteins with poor solubility. For this reason we sell a range of bacterial promoters with different expression levels (OXB1(low)>OXB20(high)) that can be provided with the peptide tags in this plasmid on request.
About the Peptide Tag:This plasmid contains an n-terminal Maltose Binding Protein (MBP) affinity tag that can be fused to a gene of interest to allow protein detection and/or purification. The sequence of the tag is: EEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEEKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLIAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGETAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFWYAVRTAVINAASGRQTVDEALKDAQTNSSS. This plasmid also contains a secondary Hexa-Histidine (6His) protein tag. The sequence of this tag is: HHHHHHWe provide a range of dual peptide tag plasmids. This is because some peptide tags provide specific biological properties (e. g., small molecule affinity new epitopes solubility or protein secretion) that are not provided by others.
About the Cleavage Tag: This plasmid also encodes a protease cleavage site that is designed to be positioned between your gene of interest and the tag to allow the removal of the tag following protein purification or isolation. This plasmid contains a Thrombin cleavage tag. The protein sequence of the cleavage tag is: LVPRGS. It cleaves preferentially between the Arg and Gly residues. Off target cleavage can often occur at non-specific sites normally from other contaminating proteases. To ensure maximal protein integrity the enzyme reagent must be highly pure.
Promoter Expression Level: This plasmid contains a constitutive bacterial promoter that does not require induction. It is the strongest bacterial promoter we sell and this can cause solubility and expression problems with some proteins. We also offer a range of other bacterial promoters that are compatible with this plasmid and are available on request.
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