Parasitic Fitness Of Blumeria Graminis F. Sp. Iritici With Different Sensitivity To Temperature And Cloning And Expression Of Heat Shock Protein Gene, HSP70

Wheat powdery mildew, caused by Blumeria graminis f. sp. tritici, a kind of strictly obligateparasite, is one of the most important wheat diseases in China. Temperature is considered as one of themost important climatic factor which affects its occurrence and epidemic. The results of recent studies,showed that temperature stress also have impacts on the pathogen population, which made changes onthe sensitivity to temperature. This change may affect the over-summering and over-wintering ranges ofwheat powdery mildew, and then affect its occurrence and epidemic degree. Research of the parasiticfitness and heat-resistant molecular mechanism are based on the B. graminis f. sp. tritici adaptation totemperature. This study will provide the basis for the delineation of the over-summering range andprediction of the disease long-term development trend under the condition of global climate change. Themajor results were summarized as follows:1. Four epidemic components: latent period, infection probability,lesion cumulative sporulationand lesion daily-expansion area of B. graminis f. sp. tritici were tested by detached leaf segment methodunder18,19,20,21,22,23℃temperature separately, and their parasitic fitness were calculated. Theresults showed that the isolates with low sensitivity to temperature have shorter latent period, higherinfection probability, larger lesion daily-expansion area, more lesion cumulative sporulation and higherparasitic fitness compared with high isolates under high temperature conditions. Furthermore, weestablished the models of the relationship between temperature (X) and parasitic fitness (Y)of B.graminis f. sp. tritici and found that Y=(28.4-X)^3.27,Y=0.81(24.8-X)^3.27 and Y=0.798(23.7-X)^2.457 werequiet fit to the isolates of B. graminis f. sp. tritici with low sensitive, moderately sensitive and highlysensitive to temperature, respectively.2. The cDNA sequence of HSP70gene was cloned using RT-PCR and RACE(GenBank accessionnumber: JQ917466). The full cDNA length was2216bp, encoding648amino acids, of which the3' endwas rich in AT-element (ATTTA)and polyA tail. Besides, the amino acid sequence contained threesignature sequences of highly conserved of the HSP70proteins family, and it had a high homology withother fungi amino acid sequences in the NCBI. Meanwhile, HSP70protein had a high conservative inthe N-terminal, but the conservation reduced at C-terminal, which inferred that its C-terminal structuremay have a larger variation in the process of biological evolution3. The research system of HSP70relative transcript levels of B. graminis f. sp. tritici wasestablished on basis of double standard curve method of real-time PCR with18SrRNA as its referencegene. The result of HSP70expression of B. graminis f. sp. tritici under the treatments of heat shock(28℃)after different times showed that mRNA expression of HSP70gene was significantly up-regulatedafter heat shock(28℃)30mins(P=0.05), reached the maximum after60minutes, and then decreased.It was inferred that HSP70gene might play an important role in B. graminis f. sp. tritici to hightemperature stress. The result of HSP70expression of B. graminis f. sp. tritici with different sensitivityto temperature showed that there are great differences of the HSP70gene mRNA expression betweenisolates with different sensitivity to temperature after heat shock(28℃)60mins. The isolates with low sensitivity to temperature have the most HSP70expression, while the isolates with high sensitivity totemperature have the least. It was inferred that HSP70gene might play an important role in differentresistances to high temperature of B. graminis f. sp. tritici with different sensitivity to temperature.