Studies On Transferring Of Drought-related Genes By Microprojectile Bombardment And Pollen Tube Pathway

Maize is an important food and feed crops in our country,it plays an increasingly important role in agricultural production.Abiotic stress including low temperature,drought,salinity and others negatively influences the quality and yield of maize seriously,of which drought is one of the most important abiotic stress factors.According to statistics data,about 2/3 of corn acreage are dry cultivation in China.Use transgenic technology,building a system for genetic transformation,introducing the drought-related genes into elite maize inbred line,and engineering drought resistant corn,to meet the rising social demand.Protein kinases are a kind of enzymes which depend on intracellular messengers,play an important role of intermediary and amplification in protein phosphorylation,and help to complete the signal transduction.They play an important role in plant stress signal transduction.It is found that many of protein Kinase families have been involved in the regulation of plant drought,high salt and low temperature signal transduction.Three protein kinases of Zm CIPK2,Zm Pti1 and Zm Pti1-1 were demonstrated that they are induced to express in maize by abiotic stress in our laboratory,and Os07G0622000 and AK072466 are from rice protein kinas family.It had confirmed that they are closely related to plant resistance to stress.In this study,we had introduced these protein kinase genes into elite maize inbred line of Ji 444,501 and J 24 by either microprojectile bombardment or pollen tube pathway.The main results were showed as the following:1.p CI-AK072466-hpt and p CI-Os07G0622000-hpt were introduced into immature embryos and callus from elite maize inbred line of 501 by microprojectile bombardment.After resistance screening,differentiation,regeneration,transplantation,detection,we proved that the immature embryos were more suitable receptor material for particle bombardment.15 and 2 transgenic lines were obtained with immature embryo and callus of 501 as receptors respectively,and transforming rate was 1.67%,0.19%,respectively.2.Zm CIPK2,Zm Pti1 and Zm Pti1-1 genes were introduced into maize inbred lines of Ji 444 by pollen tube pathway.Transforming rate was 1.33% by PCR analysis.Through the analysis of yield per plant and 1000-grains weight from T1 transgenic plants,the primary result showed that transgenic plants had improved drought tolerance comparing to control plants.3.p CI-Os07G0622000-hpt was introduced into Maize inbred lines of Ji 444 by the pollen tube pathway,the T1 generation transgenic trefoil stage seedlings were treated with the PEG-6000 which could simulate drought stress,by measuring the relative water content and electrolyte leakage of cell in leaves,the primar result showed that transgenic plants had improved drought tolerance comparing to control plants.4.Constructed the plant expression vector of p Green0229-Os07G0622000-bar,and transformed into J24 inbred line by the pollen tube pathway,finally we obtained four transgenic plants with herbicide –screening and PCR analysis.