The Race Identification,Seeds Purification Of Orobanche Cumana And Mining Of Sunflower Resistance Gene Against Race G

Orobanche cumana is a parasitic seed plant that parasitizes the roots of sunflowers in a holoparasitic manner.O.cumana can lead to a significant decline in the yield and quality of sunflower becoming a bottleneck in the development of sunflower industry besides Sclerotinia sclerotiorum.In this paper,the physiological races of O.cumana collected from seven sites in the northern foothills of Yinshan Mountain in Inner Mongolia were identified.The internationally accepted method for host identification of broomrape was used.We selected the seeds of races E,F and G,and carried out single-plant purification and propagation on the identified hosts,respectively and used the purified race G seeds to identify the resistance levels of 15 sunflower cultivars.On mining the disease resistance gene for race G,we selected sunflower lines with close lineage but different resistance levels for transcriptome sequencing and analysis.The specific research results were as follows:1.The identification results of physiological race of broomrape showed that,the O.cumana sample from Wuchuan Touhao village(THC)in Hohhot was identified as race E,and the O.cumana samples came from Huhhot Wuchuan Doudoupu village(DDP),Ulanchabu Siziwang Banner Hongqitan(HQT),Ulanchabu Siziwang Banner Jushui Town(JSZ),Ulanchabu Siziwang Banner East No.8 Township Xiaocao Nianwa(CNW)),Ulatai(WLT)of the Siziwang Banner of Ulanchabu,Siziwang Banner of Ulanchabu(SBT)were all identified as race G.2.Under greenhouse conditions,we used the internationally accepted host identification method to purify the races E and G identified above and the mixed samples of the race F seeds identified earlier in the laboratory.Finally,we obtained 17 race E,3race F,and 81 race G purified from a single plant.3.The results of the resistance identification of different sunflower varieties in the field showed that there were 5 immune varieties,3 high-resistant varieties,1medium-resistant variety,2 susceptible varieties,and 4 high-susceptible varieties in the15 tested varieties.4.We used a single purified race G to inoculate the anti/infectious Sandaomei strain for transcriptome sequencing,cleaned and analyzed the obtained sequencing data.At the same time,we performed GO and KEGG enrichment analysis on the deferentially expressed genes of resistant and susceptible strains in the large tubercle stage and small tubercle stage of O.cumana parasitism,respectively.The results showed that reactive oxygen species related genes and RAS signal pathway related genes played an important role in sunflower response to the infection of broomrape race G.By cleaning the deferentially expressed genes,13 down-regulated genes and 2 up-regulated genes were obtained in different parasitic stages.There were 15 deferentially expressed genes screened for the tubercle stage of O.cumana parasitic tubercles,of which 6 genes were down-regulated and 9 genes up-regulated.5.Taking 10 candidate genes from the above deferentially expressed genes,we performed QRT-PCR verification and found that the expression patterns of the six selected candidate disease resistance genes were in accordance with the results predicted by RNA-Seq through comparison.Therefore,these genes can be used as resistance candidate genes for functional verification.Subsequently,we also analyzed the transcripts of disease resistance related genes in resistant / susceptible sandaomei strain by QRT-PCR.The results showed that the relative expression of disease resistance related genes in resistant sandaomei strain were significantly higher than that in susceptible sandaomei strain.