Establishment Of Regeneration System From Maize Node And Study On AGPase Genes Transformation Into Maize Mediated By Agrobacterium Tumefaciens

Maize （Zea mays L.）, which is the annually gramineous plant, is one of the mostimportant grain in the world. It is widely used as feed, industrial raw materials. Maizestarch is the main component of the maize kernels. Compared with the developedcountries, maize starch research in China has a large gap, particularly in the geneticsbreedings and processing utilizations. Therefore it is particularly necessary tocultivate high-starch maize germplasm using the genetic engineering combined withthe traditional breeding method.In the transgenetic system, successful genetic transformation depends primarilyon the excellent acceptor system. The regeneration system was established from theexplants of eight maize inbred lines nodes. Compared with the callus regenerationsystem, this system could regenerate plantlets in a short period of time with theadvantages of no genotype and seasonal restrictions. The nodes of Zheng58weretransformed AGPase gene via Agrobacterium tumefaciens, and the plantlets wereregenerated, which were opening up a new way of maize genetic transformationacceptor system. The main research content and results were as follows:1. The plant binary vector harboring ZmAGPase large and small subunits geneswas successfully constructed. The T-DNA region of the plasmid contained twoobjective genes ZmAGPL and ZmAGPS, which were promoted by theendosperm-specific promoters RP5and HorD respectively.2. A regeneration system was established. The nodes of Zheng58were used asexplants to study the hormone concentration, seeding length, node size on the receptorsystem. The results showed that: the regeneration rate was highest when theconcentration of NAA in the rooting medium was0.3-0.5mg L-1, the length of theseedings was4.5-5cm, the upper and lower length of the node was0.5cm. The nodes of eight inbred lines were regenerated, and the regeneration rate was92%at the least.3. The node of Zheng58was histological observed using the method of paraffinsections. It is observed that the meristem and leaf primordias were in the node.4. The ZmAGPase genes was transfered into maize nodes mediated byAgrobacterium tumefaciens, then the nodes were selected on the medium containingthe concentration of bialaphos for3mg L^-1. Four T0transgenic plants of ZmAGPLgene were gained which were confirmed by PCR and PCR-Southern, and thedetection rate was6.06%.