Establishment Of Genetic Transformation System About Maize Shoot Apical And Genetic Transformation Of (E)-β-farnesene Synthase Gene

The corn is one of the most important food crops,playing an important role in the natio nnal economy.Aphids are major agricultural pests,a big threat to maize yield.The corn will suffer from the harm of aphids in maize growth period.How to effectively control aphids,is an important problem in the production of corn.At present,the lack of effective aphid resis tance gene in maize germplasm,and by transgenic technology to create new maize germplas m that resistance to aphid is particularly important.（E）-β-farnesene,as the main component of most aphid alarm pheromone,has significant repelling effect on aphid and attracts natural enemies of maize herbivores.The（E）-β-farnesene synthase which converts farnesyl diphosphate（FPP）to the sesquiterpene EβF has been isolated and characterized from many plants.Here,the isolation and cloning of the（E）-β-farnesene synthase gene from black peppermint was reported.Plant expression vector p FGC5941-EβF was constructed and transformed into maize using the maize shoot apical growing point as receptors.The results are indicated as following:（1）（E）-β-farnesene synthase gene was isolated from black peppermint by RT-PCR.The plant expression vector p FGC5941-EβF was obtained by ligation of EβF and fragment of p FGC5941.（2）The Agrobacterium of containing p FGC5941-EβF plant expression vector was used for transformation in maize shoot apical growing point.The results indicated that good transformation efficiency could be obtained when the concentration of bacterium OD600 was 0.6,the co-cultivation time was 2h,the vacuum infiltration time was 10 min and the receptor is maize shoot apical growing point.（3）The transformation efficiency is different when use different material.5 inbred line（zheng 58,chang7-2,qi319,zheng22,87-1）seeds and 1 cenospecies（zhengdan958）seed were used in the study.The PCR detection of T0 putative transformant show that zhengdan958 has the highest transformation efficiency 5.36%,chang7-2 has the lowest transformation efficiency0.91%,and the average transformation efficiency is 3.07%.（4）Using 3 different methods to detect the positive rate.Herbicide-resistant test in the T₀ plantlets,the positive rate is 2.96%.GUS gene expression test in the T₁ immature embryos,the positive rate is 2.1%.PCR detection of T₁ putative transformant,the positive rate is 1.91%.（5）The competent cell is a necessary condition of agrobacterium-mediated transformation.The innovation point of this research is that choosing the special stage seed as transformation receptor.（6）In this experiment,maize shoot apical growing point was choosed as receptors,the（E）-β-farnesene synthase gene was transformed into maize by Agrobacterium tumefaciens mediated transformation pathway.And a new way to create new maize germplasm resistance to aphid was explored.