Isolation Of ZmBT1and ZmAATP Genes And Identification Of ShYS, BRi Transgenic Maize

Maize is one of the most important food crops in the world, and70%~80%percent of itsgrain weight is starch. The increasement of starch content in endosperm of maize usingtransgenic technology will be helpful to improve the yield.In crops, ADP-Glucose that is the precursor of starch synthesis generated in thecytoplasm is involved in starch synthesis in plastid by transmembrane transportation. It caneffectively improves starch synthesis efficiency to increase the transport capacity ofADP-Glucose into the plastid. In this study, we cloned the ZmBT1gene encodingADP-glucose transporter protein and the ZmAATP gene encoding ATP-ADP translocatorprotein from maize. Analysis showed the ZmBT1protein belonges to the nucleotidetranslocator protein and ZmBT1gene is expressed constitutively in maize plants. TheZmAATP protein is closer phylogenetic relationship with ATPase. The expression levels ofZmAATP gene gradually increase in the maize grain after pollination. The expression cassettesof ZmBT1and ZmAATP genes drove by the constitutive promoter (Ubi) or theendosperm-specific promoter (GluAI) were constructed respectively.In this study, we successfully introduced transferred modified starch synthesis ofADP-glucose pyrophosphorylase gene (ShYS) and starch branching enzyme RNAinterference genetic transformation vector (BRi) into the Qi319inbred usingAgrobacterium-mediated transformation system. We obtained six hundred and sevenresistant plants out of14,661explants. We have obtained170seed setting transgenic linesfrom the T0generation. One hundred and three transgenic lines were planted and the seedsof those lines was analyzed after harvest. Ten transgengic lines were identified by PCR, inwhich contains Ubi::BRi genotype7lines and Ubi::ShYS genotype3lines. The above lineswere further confirmed at the genomic level, protein levels and gene expression levels in theT2generation. The detection results using fluorescent quantitative PCR showed that the expression of ShYS gene in the positive plants was significantly increased, while theexpression of SBEI and SBEIIb genes was partly inhibited. ELISA results showed thatexogenous NPTII resistant protein had expressed in transgenic plants. Comprehensivemolecular detection results showed that we have successfully obtained the overexpressionUbi::ShYS and the RNA interference UBI::BRi of transgenic plants.