The Cloning Of HSF1 Gene In Maize

Our previous data have demonstrated that maize cytosolic HSP70 protein could bind to CaM in the presence of Ca2+ and the CaM-binding site in HSP70 was identified. Following works proved that Ca2+ signaling pathway is involved in the inducement of heat shock protein, and the acquisition of the heat-shock-induced thermotolerance in wheat;In eukaryota, the express of heat-shock-gene is regulated by HSF,which is activated by binding to HSE. Among the several tomato HSFs found and studied so far, HSF1 play a central role for the heat shock response in tomato.The full-length tomato HSF1 cDNA could not be obtained by method of RT-PCR, but a 360bp of DNA fragment from HSF1 cDNA(nucleotide sequence from 480 to 840) was obtained by RT-PCR.The analysis of antigenic determinant showed that there are 5 epitopes on the amino acid sequence which was deduced from the nucleotide sequence obtained above.Zmhsfa is a HSF constitutively expressed in maize.To obtain zmhsfa cDNA in maize, a cDNA λ -ZAPII library from maize seedlings was constructed and screened with a 291bp of DNA fregment ,but faild. A PCR product encoding the DNA-binding domain of zmHSFa protein (first 118 amino acid) was obtained by RT-PCR.The result of antigenic determinant analysis revealed that the deduced amino acid sequence from the PCR product contained 4 epitopes, and could be used as antigen.The DNA fregment was constructed into expression vector pET-28b.the construction was showed to allow high level expression of maize extraneous protein in E.coli.