Endosperm-specific Expression Vector Construction And Genetic Transformation Of Wheat Avenin-like B Gene

Wheat Avenin-like protein is a new type of seed storage proteins of unknown function. The sequence of Avenin-like protein was predicted by the sequence of avenin-like gene, riching in cysteine residues. Avenin-like protein is speculated to affect processing quality of wheat flour. Currently, the relationship between Avenin-like protein and processing quality has not been reported yet.In this study, the avenin-like b gene was amplified from the genome of wheat and then was cloned into vector pLRPT to construct eukaryotic expression vector pLRPT-avel, and then was transformed into Zhengmai 9023 and Emai 12 by particle bombardment. In this research we got the main results as follows:(1) Selected the expression vector pLRPT without target gene and the vector pAHC25 with marker gene. We identified vector pLRPT by restriction and sequencing analysis. The eukaryotic expression vector pLRPT-avel with 1Dx5 promoter that expressed specificlly in the endosperm was successfully constructed, then it was transformed into wheat by particle bombardment.(2) Compared the induction mediums with different concentrations of sucrose on Zhengmai 9023 and Emai 12 of callus induction, and ultimately determined the concentration of 3% sucrose induction medium fit for the induction of Zhengmai 9023 and Emai 12.(3) The avenin-like b gene was transformed into Zhengmai 9023 and Emai 12 by particle bombardment, the transformants were confirmed by PCR to amplify the Gus gene and CAMV35S genes. The proportion of the number of positive transgenic positive in the total regeneration is nearly 10%The eukaryotic expression vector pLRPT-avel expresses specifically in the endosperm was successfully constructed, and then it was transformed into wheat by particle bombardment. The results of present research provided with new research material for further study on the relationship between avenin-like b Gene and processing quality.