Cloning And Analysis Of Three Cold Shock Proteins (CspA, CspB And CspG) In E.coli BX△AG Strain

When an exponentially growing culture of Escherichia coli is shifted from 37℃to low temperature(10-15℃), an arrest of cell-growth will occur. This period, which is called the acclimation phase, is transient. During this growth lag period, the synthesis of most cellular proteins is sharply reduced, whereas a set of at least 26 cold shock proteins which play essential roles in the resumption of cell transcription and translation are immediately induced. After approximately 2-4h, cell growth resumes.CspA family in E.coli which is reported first by Jones and his colleagues in 1987 is highly researched. Jones found that CspA family consists of nine homologous proteins which was called CspA to CspI. Four of these proteins, CspA, CspB, CspG and CspI, transiently and significantly induced upon low temperature. During acclimation phase,The four cold shock proteins are considered to regulate the cells at the level of the transcription, mRNA stability and the translation. In this paper, plasmid pET-28a(+) was used for construction of expression carrier into which the coding region of gene cspA, cspB and cspG was fused into the pET-28a(+) respectively. These three expression carrier were transformed into defective strain BX△AG which is not able to grow normally under low temperature. The growth condition of the three recombinant strains (BX△AG -pET28(+)a-cspA, BX△AG -pET28(+)a-cspB and BX△AG -pET28(+)a-cspG) and defective strain BX△AG under normal and low temperature were detected. Statistics analysis software SPSS11.5 was applied to contrast and analyze the growth-level differences among the three recombinant strains against the defective strain BX△AG. SDS-PAGE was also applied to detected the expression level under 37℃and 16℃among strains BX△AG -pET28(+)a-cspA, BX△AG -pET28(+)a-cspB and BX△AG -pET28(+)a-cspG.The results showed that strain BX△A G-pET28(+)a-cspB and BX△AG-pET28(+)a-cspA can reinitiate its cell growth under 16℃. The former had an obvious extent of growthing, the latter lesser. But strain BX△AG-pET28(+)a-cspG appeared no renewing growth ability. There were also no differences among the whole protein expression level of defective strain BX△AG and three recombinant strains.