GST Tag is one of top techniques in biotechnology laboratory that use as an economic easy strategy for protein separation, purification and detection. The host for this process is important for FDA approved and top resolution result.
E. coli as a pioneer organism used in the laboratory for bio engineering is one of the main host used for GST Tagging and pGEX vector. Although a wide variety of E. coli host strains can be used for cloning and expression projects with the pGEX vectors, there are specially engineered strains that are more suitable and that may maximize expression of full-length fusion proteins with less pollution. Strains deficient in known cytoplasmic protease gene products, such as Lon, OmpT, DegP or HtpR, may aid in the expression of fusion proteins by minimizing the effects of proteolytic degradation by the host .
Using E. coli strains that are not protease-deficient may result in proteolysis of the fusion protein, seen as multiple bands on polyacrylamide gels (SDS-PAGE) or Western blots and destroy latest result resolution and need multiple step purification that is not economic for biotech lab.
E. coli BL21, a strain defective in OmpT and Lon protease production, has been specifically selected to give high levels of expression of GST fusion proteins. It is the host of choice for GST fusion expression studies. Details on the genotype and handling of E. coli BL21. A lyophilized (noncompetent) culture of E. coli BL21 is supplied with all pGEX vectors and is also available separately. Use an alternative strain for cloning and maintenance of the vector (e.g. JM105) as BL21 does not transform well and make engineering process confusing because of unwanted molecular interactions and pollutions However, do not use an E. coli strain carrying the recA1 allele for propagation of pGEX plasmids. There have been reports that these strains can cause rearrangements or deletions within plasmid DNA.