High-resolution Mapping And Exploration Of The Brown Planthopper Resistance Candidate Genes In Two Rice

Rice（Oryza sativa L.）is not only an ideal model plant for molecular biology research of monocots,but also one of the most important food crops in China and the world.The brown planthopper,（Nilaparvata lugens St?l;BPH）,is one of the most destructive insect pests which seriously damage rice production.Exploration the resistance genes from resistance germplasm and breeding insect-resistant varieties were considered as an safe and effective strategy to control BPH.In this study,CL48and CL123 were used as the resistance germplasmthe.The genetic linkage mapping populations and genetic linkage map were constructed by crossing and backcrossing with CL48 and Kangwenqingzhan（KW）as well as CL123 and 9311,and the brown planthopper resistance genes were high-resolution mapped,and the resistance candidate genes were analyzed.Previous study indicated that the brown planthopper resistance genes were located on the 11th chromosome between 11M16.89 and 11M18.41 by using F₂populations which were constructed by crossing with CL48 and KW.In this study,we used markers to screen out 100 recombinants from 8,674 F₃plants,and the resistance genes were high-resolution mapped to a 55-Kb region between YL5 and YL12 by using F₃populations.RT-PCR results of 4 candidate genes（G1/G2/G3/G4）in the region showed that the expression levels of G2,G3 and G4 were significantly different in resistant and susceptible plants at each time point of the brown planthopper feeding.Combined with bioinformatics analysis,we took R gene G4 as the key target gene for research,than we constructed a complementary vector of G4 and obtained 16 positive T₀transgenic plants.Meanwhile,the binary expression vector p YL-Cas9-g ^G4of G4was constructed by CRISPR/Cas9 system.In addition,the resistance mechanism of CL48 against brown planthopper was comprehensively studied from antibiosis,antixenosis and tolerance.The results indicated that the resistant genes exerted antibiotic effects in CL48 plants,but it was no obvious antixenosis and tolerance effect on the brown planthopper.Previous study indicated that the resistance genes were located on chromosome4th between the markers RM3.688 and RM11.353 by using F₂populations which were crossed with CL123 and 9311.In this study,the resistance genes were further mapped to a 0.799 Mb region on chromosome 4th between the markers RM3.688 and RM4.487 by BC₄F₃populations.In addition,the statistical results of the agronomic traits of BC₅F_2:3lines showed that it was not significantly different between BC₅F_2:3lines and 9311 in the grain length,grain width and 1,000-grain weight which indicated that the genetic background of BC₅F_2:3lines were highly similar to 9311.This study laid a foundation for the location and cloning of resistance genes as well as the resistance mechanism of BPH.