Transformation Of Gene MiR396 Into Wheat Varieties By Microprojectile Bombardment And Agrobacterium Tumefaciens Infection

Wheat(Triticum aestivum L.) is one of the important cereal crop in global, and no one deny the importance of this crop. Although the genotype of this crop little bite difficulty to study as compared other crops, Great progress of applying transformation technology to improve wheat characters such as productivity, quality, resistances to diseases, insects and adverse conditions have been made in recent years. Every year there are some articles which talk about genetically modified related to stress-tolerance. The study in wheat about miRNAs is less reported. It becomes a research focus in recent years, and his works haven’t been sufficient studies. There are some articles reported that the potentials of miR396 for use in genetic modifications of the important agronomic traits of plant development, abiotic stress response, and /or flowering time control. It may shortened internodes, increased tillers. MiR396 is a conserved small RNA which is also involved in multiple stress responses, however, experimental functional studies are lacking. In this study, the foreign gene miR396 was transferred into wheat immature embryos by microprojectile bombardment- and Agrobacterium tumefaciens mediated, and the factors influencing wheat transformation were analyzed. We are looking forward to obtain transgenic plants. the results were follows:(1)Transformation of wheat with miR396 gene mediated by particle bombardment and molecular identification:In this study, miR396 gene was introduced into five wheat varieties, Huamai2566, Huamai2152, Zhengmai9023, Kenong199, Longchun23,by particle bombardment, the purpose of this study is to acquire transgenic wheat with target genes, then we got it,PCR preliminary detection results showed that 7 plants showed positive. Including 5 plants of Kenong199 and 2 plants of Longchun23.We don’t know if the foreign gene miR396 had already integrated into genome of receptor plants.(2) Transformation of wheat with miR396 gene mediated by by Agrobacterium tumefaciens and molecular identification:We transformed miR396 into wheat immature embryos by agrobacterium-mediated in establishing that transformation system, Based on research with 4 different wheat varieties and the tetraploid durum wheat LDN, we only obtain 2 LDN positive plants, there is no positive plants in common wheat.(3) phenotype investigation about the positive plantsIn this study, the 2 LDN positive plants we obtained by Agrobacterium tumefaciens, theyhave no seeds, only heading. Since there are less positive plants, we are not sure if the miR396 gene regulate the internode length and tillering number.