| HrpZ is a proteinaceous bacterial elitor from Pseudomonassyringae pv.Syringae, inducing a hypersensi-tive response(HR)in non-host plants,Characterized by the rapid,localized death of plant cells at the site of pat-hogen invasion.The HR can prevent further multiplication and restrict the spread of the pathogen. It was found that activation of other defencerelated responses often accompanies HR,for example,the oxidative burst,the production of antimicrobial compounds,and enzymes involved in the general pHenylpropanoid pathway. The nonlocalized and long term induced protection,is similar to systematic acquired resistance (SAR).Maize(Zea may L.)is one of the most important crops,staple food in the world. it was also a modern food industry and an important raw material of medical and chemical industry.It is the best-studied and most tract-able genetic system among the cereals,making it the premier model system for studying this important group of crops. Highly efficient regeneration system is always the focus of scientist researches.Maize yield in China is next below that in the United State. However,huge yield loss and deteriotation of quality of cultivated plants were caused due to various diseases.The crop breeding of disease resistance is limited because of lacking the resistance resource and the hereditary isolation of different plant species. Plant gene transformation can solve the proplem to improve genetic melioration and enhance resistance to various diseases and stress of maize by gene engineering technology.The current advancement of biotechnoloy has ushered agricultural reasearch into a new ear of transgenic organisms.which plays an important role in national economy.In order to select and establish the regeneration and transformation system of maize elite inbred lines, we systematically explored the characteristics and parameters of maize elite inbred lines callus inducement which induced by immature embryo, stem apex,root apex and mature embryo as explants, researched the factors which affect the induction,differentiation and regeneration and the sytem of Agrobaetirium tumefaeiens-mediated transformation.The integration of targeted gene in transgenic plants was proved by PCR.The main results are summarized as follow:1. A total of ten widely cultivated maize elite inbred lines were researehed on callus inducement and regeneration abilities. The induction of callus and regeneration were optimized in this study and genetic breeding research of maize can get an important technology support from this system.Primary callus were eas-ily induced in ten maize cultivars, but the frequencies of embryogenesis callus were significantly different. The results showed maize regeneration abilities were highly genotype-dependent.The embryogenesis callus were induced in six maize elite inbred lines,which were H99,Dan988,Tie7922,1322,7319 and PH4CV.Besides the callus inducement were studied with immature embryo,mature embryo, stem apex,root apex and spire of maize inbred lines (H99,Dan 988 and Tie7922) which were sifted for years in our lab as explants.The results showed that primary callus could be induced from root apex and spire,Only Dan 988 can be induced callus from immature embryo.mature embryo,stem apex in three genotypes,so that we can know Dan 988 was quality variety for callus induction. Suitable concentration of 2,4-D and 6-BA could improve the induction percentage of stem tip callus,also kinds of medium,embryo age,freezing time at 4℃.different addition like KT,CaCl2 and AgNO3 have consequences for callus from immature embryo.2. In this study, Through orthogonal design,immature embryo callus from stem tip of maize inbred lines Dan 988 was infected by A. tumefaciens strains with vector pCAMBIA1301-HrpZpsta and pCAMBIA3301-HrpZpsta.The factors like Agrobacterium tumefaciens concentration and incubation-time, concentration of acetosyringone(AS) and co-culture time which influencing transformation efficiency were studied and optimized.The highest transformation rate was found with the optimized combination of 0.6 OD×30 min for Agrobacterium tumefaciens concentration and incubation-time,2 days for co-culture time and 200μmol/L for concentration of acetosyringone. Besides the sensitive characteristic on saline-alkaline and resistance to glypHosate of maize callus(H99 and Dan 988)was studied. determine the critical concentration on saline-alkaline and resistance to glypHosate.Callus come of stem tip the rate of resistance callus, regeneration rate and percent conversion all a little more than immature embryo,and higher thanmature embryo.The recombinant plasmid with HrpZ gene was transformed inio maize callus with Agrobacterium tumefadensme diated. It was proved that HrpZ gene had been integrated into the transgenic maize genome and was expressed at transcription level by PCR,18 plants were embedded target gene HrpZ resistance to salt and 10 plant resistance to glypHosate of H99,12 plant resistance to salt and 5 plant resistance to glypHosate of Dan 988.But not obtain T1 progeny fruit.3. In the study, we discussed DNA concentration and the time of DNA heeded by pollen tube pathway, only Dan 988 obtain two transgenic plants. Although the conversion efficiency of pollen tube channel method is lower, but it can avoid complications of tissue culture, difficulty of regeneration plant transplanting survive, it still a kind of good method.
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