Wheat Genetic Transformation Via The Pollen-tube Pathway And The Analysis Of Gliadin In Mutant

Wheat is one of the most important foodstuff crops in China. The wheat breeding methods include particle bombardment transformation, Agrobacterium-mediated transformation pollen-tube pathway transformation in common use. Pollen-tube pathway transformation has been being adopted broadly for its simple operation and higher transformation rate, so far, many wheat individuals with adversity-resisted genes have been breeded by this method.In this study, in order to breed wheat with better drought tolerance, wheat is transformed by non-emasculation pollen-tube pathway transformation and artificial emasculation pollen-tube pathway transformation, with donors DNA from oat, hexaploid triticale, millet, maize, wheat with good drought tolerance and plasmid pBI1301.1. The establishment of transformation system. The experiment materials of 2005 were transformed by non-emasculation pollen-tube pathway transformation, their average setting percentage was 43.9% and average percentage of survival was 56.0%; the experiment materials of 2006 were transformed by artificial emasculation pollen-tube pathway transformation,their average setting percentage was 65.5% and average percentage of surviva was 73.6%. Plasmid pBI1301 was transformed into receptor wheat 1145 by non-emasculation transformation. The DNA was extracted from the leaf of the 18 individuals seedlings in M₀ for PCR testing. The result showed that No.15 individual was positive, the seedlings in the following generation were tested by PCR and histochemical pigmentation for GUS, the results showed that the mudi DNA fragment were not amplified and there was not transient GUS expression. The results of tests showed that inserted DNA fragment form plasmid pBI 1301 was not inherited from M₀ to M₁. In 2006, transformed by emasculation and detected by histochemical pigmentation for GUS, among the transforming groups with plasmid pBI 1301 donor, the expression of the GUS was tested in three transformed strain : No.12-2(Fan 1/1301), No.25-3(Fan 3/1301), No.36-3(Wen 19/millet+1301), the result show that the blue splashes appeared in each strain. Furthermore, the DNA of these three individuals leaves were tested by PCR, the result showed that the GUS was amplified in these three individuals. In a word, the artificial emasculation transformation is better than non-emasculation transformation, it has the distinct advantages.2. Analysis of gliadin in mutant. A mutant 406A was obtained from 1230 transformed by artificial emasculation pollen-tube pathway transformation with the total DNA from Yuanfeng998, its harvest period was 3 days longer than the strain 1230. Other agronomic traits were the same as 1230. The gliadin in seeds of M₀ were tested by A-PAGE, the results showed that the gliadin bands of some seeds were different from that of 1230, it suggested that the transformation of exogenous DNA had made the variations of gliadin of the wheat.