Cloning And Prokaryotic Expression Of Avenin-like B From Wheat Variety Zheng 9023

Wheat grain quality is defined as the suitability of end-use of wheat, including nutritional quality and processing quality. Processing quality of wheat is still relatively inferior in China, but the demand of excellent processing quality of wheat is increasing, so the improvement of wheat quality is an important goal of wheat breeding. It is an effective way to do this through the use of conventional breeding and transgenic technology.Wheat avenin-like gene is a newly discovered type of seed storage protein gene, which is divided into avenin-like type a and avenin-like type b. There are no relevant reports about the relationship between Avenin-like protein and wheat quality. avenin-like genes were isolated from Zheng 9023 and the antibody was prepared. We attempt to analyze the basis for the Avenin-like b protein integrating into gluten protein aggregates. The main results are as follows:(1) Four novel avenin-like b gene's ORFs were isolated from from the wheat genome of Zheng 9023. These ORFs are 855 bp in length. The deduced protein sequences show that each contains 19 cysteine residues. There is typical "Cys-Cys "domain. The first 1-18 N-terminal amino acids of each deduced protein are predicted to be a signal sequence.(2) The avenin-like b gene was amplified and then was cloned into vector pET32a to construct prokaryotic expression vector pET32a-avelb. Avenin-like b protein was successfully expressed in Escherichia coli BL21. The purified protein was used to immunize New Zealand rabbits and polyclonal antibody was raised.(3) Avenin-like b protein from Zheng 9023 was detected by Western blot, which lays the foundations of analyzing further Avenin-like b protein involving in the formation of gluten protein aggregates.