| Stripe rust , caused by puccinia striiformis f.sp.tritici, is one of the most serious diseases of wheat in the worldwide, and planting resistant cultivar is the most effective and economical method to control this disease. Many researches have been carried out on different aspects of wheat stripe rust, such as from morphology,histology,cytology,physiology biochemistry. However, studies on the molecular mechanisms about host resistance have attracted relatively little attention. In order to find the characteristics of PRS gene transcript accumulation pattern in incompatible and compatible interactions between wheat and puccinia striiforms, to understand the relationship between defense response genes transcript accumulation characteristic and host resistance. Recently, we examined defense response genes transcript accumulation pattern by Northern blotting. We achieved one new gene from wheat leaves by 5'RACE and 3'RACE, it's transcript accumulation pattern was similar to PR-2(?-1,3-glucanase). This research had provided a theoretical basis to find the molecular mechanisms to wheat resistance. Consequently, the research described in this thesis focused on the above two aspects. 1. A simple and rapid method of extracting total RNA from wheat leaves was proposed .The method was also especially useful for total RNA extraction of plant material with plenty of polysaccharide and lipids. It can be used for RT-PCR, RACE, Northern blotting. 2. PR-1, PR-2(?-1, 3-glucanase), PR-3(chitinase), PR-5(thaumatin) defense response gene transcript accumulation pattern in incompatible and compatible interaction in wheat leaves following puccinia striiforms. (1) PR-1 defense response gene transcript accumulation pattern was: In incompatible interaction, it showed detectable accumulation at 24 h after inoculation, there was one peak at 48 h, there had higher level accumulation between 30-48 h, there still had accumulation at the later stages following inoculation. In compatible interaction, it was not detected until 42 h after inoculation. It can be observed that it occurred earlier and had higher level accumulation in the infected incompatible wheat leaves than in the compatible one. (2) PR-2(?-1, 3-glucanase) defense response gene transcript accumulation pattern was: In incompatible interaction, it could be examined at 18 h after inoculation, there was higher accumulation at 30 h, and there was more accumulation level at the earlier stages than at the later stages after inoculation. In compatible interaction, the transcript accumulation pattern was similar to the incompatible one, but the accumulation level was lower than in incompatible. It was not found in control. (3) PR-3(chitinase) defense response gene transcript accumulation pattern was: In incompatible interaction, it was occurred at 12 h. There was one peak at 30 h, from 30-48 h, and there was much accumulation. There was also higher accumulation at the later stages after inoculation. In compatible interaction, it was detected at 18 h, the accumulation peak was at 42 h. It had strong late accumulation at the later stages than at the earlier stages after inoculation. There was detectable accumulation in control. A somewhat earlier and higher accumulation in the incompatible interaction than in the compatible one. The accumulation peaks occurred early in incompatible than in compatible. There was higher accumulation in incompatible interaction than in compatible one, but it was contrary at later stages after inoculation. (4) PR-5(thaumatin) defense response gene transcript accumulation pattern was: In incompatible interaction, it was expressed at 12 h, the peak was at 30 h. In compatible interaction, it was similar to incompatible interaction one. There was no detectable accumulation in control. At the indicated time points after inoculation, there were higher accumulations at later stages than at the earlier stages both in incompatible and in compatible interaction. 3. PR-1, PR-2(?-1, 3-glucanase), PR-3(chitinase), PR-5(thaumatin) defense response genes were induced by puccinia striiforms. They were none specific resistance genes. It was clearly that they occurred earlier and had higher accumulation in incompatible interaction than in compatible interaction. 4. It is clear that induction of PR-1, PR-2(?-1, 3-glucanase), PR-3(chitinase), PR-5(thaumatin) defense response gene in wheat leaves did not afford completed protection from the invading puccinia striiforms, other factors such as the speed of defense response gene production, the level of accumulation, species of the invading puccinia striiforms, they also play important roles in host resistance. 5. 5'RACE and 3'RACE (Rapid Amplification of cDNA Ends) have been used to clone one new ?-1,3-glucanase gene in wheat leaves infected by puccinia striiforms, it was submitted to GenBank, the GenBank accession number is DQ090946. It full length is 1338bp, it has been found out that it has the complete open reading frame and is classified as 334amino acids with codes. It is coincide with KOZAK principle. Amino acid multigene alignment analysis showed that the amino acids have the higher identity with other plant; the highest percentage is 95.8%. The Northern blotting pattern was the similar to PR-2( pBH72-I1 ?-1,3-glucanase). |
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