Population Genetic Diversity Of Blumeria Graminis F.sp.tritici In China And Expression Level Of Calreticulin Gene In High Temperature Tolerant Isolates

Wheat powdery mildew caused by Blumeria graminis f.sp.tritici(Bgt),an obligate parasitic fungus,is typical airborne disease and currently one of important wheat diseases in China.Under the background of global warming,the high temperature tolerant isolates of Bgt were isolated in the fields.(?)To determin the genetic relationship and gene flow of Bgt populatios among 16 provinces(cities)in China,the virulent and genetic diversity of Bgt isolates were tested and analyzed.(?)To understand the mechanisms of high temperature tolerance of Bgt,the temperature-sensitive and temperaturetolerant isolates were screened to measure the expression level of calreticulin gene.These studies provide the basis for forcesting and controlling of wheat powdery mildew.(1)The genetic diversity from the molecular level of 434 Bgt isolates ifrom 16 provinces(cities)was assayed by using SSR.The results showed that,The polymorphic loci of genetic diversity was the highest in Shandong and the lowest in Xizang.The Nei index of genetic diversity of isolates in Hebei was the highest followed by the isolates in Henan,but the lowest was in Xizang.While the Shannon's information index of the isolates was hightest in Shandong followed by Hebei,and was lowest in Xizang.Six dendrogram groups were classified based on genetic diatance.One group included Sichuan,Gansu and Qinghai.The other group included Anhui,Hebei,Henan,Beijing,Guizhou,Jiangsu,Hubei,Shaanxi and Shandong.Xinjiang,Xizang,Yunnan and Zhejiang were respectively independent group.The genetic distance of Bgt was the closest between Hebei and Henan.And the genetic distance of Bgt was the farthest between Xizang and Yunnan.There was a significant positive correlation between the genetic distance and the geographic distance.It indicated there was gene flow between the 16 provinces(cities).(2)Virulence of 434 Bgt isolates from 16 provinces(cities)was tested by using 29 cultivars/lines with known Pm genes.This study showed that:(?)Virulence frequencies of Bgt isolates ranged from 80% to 100% to Pm3 c,Pm3f,Pm3 e,Pm5a,Pm6,Pm7,Pm8 and Pm19.It suggested resistant genes are almost invalid and unsuitable in wheat breeding.Virulence frequencies of Bgt isolates was under 30% to Pm12,Pm13,Pm16,Pm21 and Pm35.Less than 5% of the 434 isolates were virulent to Pm12 and all of them was invirulent to Pm21.These results suggested these genes were excellent resistant genes in breeding,especially Pm12 and Pm21.Virulence frequencies of Bgt were different in most provinces(cities)to Pm24,PmXBD,Pm46 and so on.So these genes should have adequate distribution in agricultural production.(?)The mean of virulence genes number were different in 16 provinces(cities).The most genes number was detected in Zhejiang following Jiangsu and the least genes number was detected in Guizhou.The polymorphic loci of virulence diversity in Shandong was the hightest,and the lowest in Xinjiang.The virulence gene diversity in Hebei was the highest,but it was the lowest in Xinjiang.Dendrogram of virulent distance of 16 province(cities)had three groups.One group included Qinghai,Sichuan,Beijing,Gansu,Yunnan,Henan,Hebei and Anhui,and one group included Jiangsu and Zhejiang,and the other group included Shandong,Hubei,Shaanxi,Guizhou,Xinjiang and Xizang.The virulent distance of between the Hubei and the Shaanxi were the least.But there were the farthest virulent distance between the Xinjiang and the Zhejiang.There was a significant positive correlation between the virulent distance and the geographic distance.It indicated the virulence was not only related to the host,also related to gene flow between the 16 provinces(cities).(?)The correlation analysis between the genetic diversity and virulent diversity of Bgt isolates indicated that theywere positively correlated in the three parameters,included the observed number of alleles,effective number of alleles and the polymorphic loci.But there is no significant correlation in two parameters,included the Nei index and the Shannon's Information index.(3)The analysis of gene DNA and cDNA sequences of calreticulin in Bgt showed that the full length of calreticulin gene DNA and cDNA sequences,respectively,were 1887 bp and 1650 bp.Its cDNA sequence was a complete open reading frame encoding 549 amino acids.Meanwhile,21 of the 549 amino acids were the signal peptid sites,from 1 to 21.No transmembrane domains were predicted in the calreticulin.However,the calreticulin had conserved domain of calreticulin family.Temperature sensitivity of Bgt were verified to screene the three sensitive isolates(13-1-5-5-1-(2),13-10-5-3-(1)and 13-10-5-1-(1))and three high temperature tolerant isolates(13-14-1-(2),13-14-8-2-(2)and 13-10-5-3-(2)).The expression levels of calreticulin gene of temperature-sensitive and temperature-tolerant isolates were measured by treatment in different time at 24?conditions.The expression levels of all isolates with the high temperature treatment,whether they were temperature sensitibility and temperature tolerance,were higher than those without the high temperature treatment.Although the calreticulin gene of the isolates with different treatments mainly had two peaks in the expression levels.The expression levels of the different isolates with the same treatment had significant difference.The sensitive isolate 13-1-5-5-1-(2)with high temperature treatment in 1/2h,6h and 8h and sensitive isolate 13-10-5-1-(1)which high temperature treatment in 2h had higher expression levels than other isolates.But the expression levels of temperature tolerant isolates were higher than temperature-sensitive isolates with other treatments.