Receptor Selection From Wheat Mature Embyos, Introduction Of Chitinase Gene In Wheat Var Particle Bombardment And Study Of The Insert Point

Wheat (Triticum asetivum L) is one of the most important food crops in the world and also one of which has the widest planting area in our country. The production and quality of wheat play a very important role in the development of our economy and the raise of living standard of the people. But the production and qualities are affected by the weather, disease and pests. The key point of the wheat breeding is to find a excellent breed has good production and high quality. Plant genetic engineering technology provides a new method for wheat breeding to quicken the breeding procedures and shorten the breeding cycle. Transformation for elite wheat cultivars with good agronomic traits can find new varieties of wheat with great applicable value in a short time. And find the varieties with excellent tissue culture traits mean a lot to the development of wheat genetic engineering.In present study, mature wheat embryos without radicle of 7 different wheat genotypes MianYangll, ChuanNong20, ChuanNong21, ChuanNong23, ChuanNong24, ChuanNong25, ChuanNong26 were used as explants for culture in vitro. Two wheat genotypes were selected out as the wheat varieties with excellent tissue culture traits for they displayed a high regeneration response. Then immature embryo callus of wheat cultivars MianYang11 and ChuanNong23 were bombardmented using PDS 100/He particle delivery system transformed plasmid vector pubichitpBC1300 and pAHC-20, which contained a chitinase gene and a bar gene respectively. By PCR analysis and sequencing blast we could be sure that the chitinase gene has integrated into wheat genome and 6 regenerated wheat plants have the foreign gene were acquired.Results of the study are as following:1. The 7 different wheat genotypes have an average value of callus formation rate over 95%. But only two genotypes of MianYangll and ChuanNong23 have a high differentiation rate over 45% while there're two genotypes have a differentiation rate below 30%; and also the two wheat cultivars MianYang11 and ChuanNong23 have a high efficiency of regeneration over 30%, to the contrary, the rest five wheat cultivars only have an efficiency of regeneration lower than 25%. The efficiency of callus formation of mature embryos has no significant difference between genotypes, but the efficiency of differentiation and regeneration has. By the correlation analysis, there's no correlation between the efficiency of callus formation and differentiation, also has no correlation between the efficiency of callus formation and regeneration. But there's a high correlation between the efficiency of differentiation and regeneration. So the wheat cultivars have high efficiency of differentiation can be chosen as a cultivar has excellent tissue culture response. So the elite genotypes MianYangll and ChuanNong23 were selected for acceptor genotypes in wheat transformation.2. By the tissue culturing of immature embryos of MianYang11 and ChuanNong23, both the two genotypes immature embryos have higher percentage of callus formation, differentiation and regeneration than the mature embryos, which suggested that the chosen genotypes Mian Yang 11 and ChuanNong23 have good tissue culture response.3. Ttransform chitinase gene and bar gene into MianYangll and ChuanNong23 by particle bombardment, then 6 regeneration wheat plants were acquired.4. We have got six electropositive wheat regenerated plants by the PCR analysis, and after sequencing blast we can conclude the gene has been transferred into these six regenerated plants.5. According to the electropositive regenerated plants, MianYangll has a transformation frequency of 0.45% and the frequency of ChuanNong23 is 0.22%.