Molecular Mapping Of Gene For Small Brown Planthopper And Rice Black-streaked Dwarf Virus Disease Resistance In Rice Variety 9194

The small brown planthopper(SBPH),Laodelphax striatellus Fallen(Homoptera:Delphacide),is one of the most destructive pests and widely spread throughout the main rice-growing areas in Asia.It causes serious damage not only by directly feeding,but also by transferring two major virus diseases,viz.rice stripe virus disease(RSVD)and rice black-streaked dwarf virus disease(RBSDVD).In recent years,RBSDVD has become the most serious disease in the rice-growing regions of middle and lower reaches of the Yangtze River and North China.Once the plants are infected with RBSDVD,it is generally incurable.In the rice production,the spray of chemical pesticides is still the main way to control SBPH and RBSDVD,but with increases in migration behavior and outbreak of SBPH,chemical control is not satisfactory.In addition,it is expensive and harmful to the environment,and it can also urge SBPH to readily develop resistance,so exploring new genes from diverse sources and developing new resistant cultivars are considered to be the most economical and effective strategy for SBPH and RBSDVD management.The disease is caused by RBSDV and transmitted by SBPH in a persistent manner but is not transmitted to offspring through the ovary,so the identification of RBSDVD became complex and difficult,and very few studies had focused on RBSDVD research.The identification and mapping of insect-or virus-resistant genes is helpful for speeding up the application in breeding program.In this study,we researched the RBSDVD and SBPH resistance of indica cultivar 9194 by natural infection in the field and artificial inoculation.The results showed that 9194 is highly resistant to RBSDVD and SBPH.Afterwards,QTL analysis was performed using a F2:3 population derived from a cross between 9194 and a susceptible japonica variety Suyunuo(SYN).And qRBSDVll,a RBSDV-resistant QTL,was further fine-mapped by BC2F2:3 population.The results of this study were as follows:1.Genetic analysis and molecular mapping of QTL for rice black-streaked dwarf virus resistance in rice variety 9194.(1)RBSDVD natural resistance evaluation was deployed in two different regions for six years(2010 一2016,except for 2014).The results showed that 9194 is a highly and stablely resistant indica cultivar coferring resistance to RBSDVD,and SYN is a highly susceptible japonica variety.The result was further confirmed by artificial inoculation identification.Thus those two varieties are ideal material for the research of RBSDVD.(2)To identify QTLs for RBSDVD resistance,a F2 population including 796 F2:3 lines derived from 9194 and SYN was used in natural infection in 2012.Among them,212 lines with well repeatability were selected to detect QTLs for RBSDVD resistance.The disease incidence(DI)frequency distribution of the F2:3 population varied continuously,ranging from 0-87.5%.It indicates multi-QTLs may ascribe to RBSDVD resistance in the population.Four QTLs for RBSDVD resistance,designated as qRBSDV3,qRBSDV6,qRBSDV9 and qRBSDV11 were detected on chromosomes 3,6,9 and 11 with LOD scores of 2.34,4.42,7.26 and 7.16,respectively.These QTLs explained 5.4%,10.3%,35.5%and 31.1%of the phenotypic variance,respectively.Jointly they explained 82.3%of total phenotypic variance in this population.Resistance alleles of the four QTLs were all derived from 9194.Among them,qRBSDV9 and qRBSDV11 were two major QTLs,while other two were minor QTLs.To introgress and pyramid RBSDVD QTLs to SYN,we firstly crossed 9194 with SYN and then resultant hybrid F1s were continuously back crossed with SYN for 3 times.Finally eight lines from the BC3F2 harboring none,single,two or three loci of qRBSDV6,qRBSDV9 and qRBSDV11 were selected to evaluate the RBSDVD resistance,these data further confirmed the above three QTLs are reliable and can act additively.(3)In this study,qRBSDV11,a RBSDV-resistant QTL,was further confirmed by a newly constructed BC2F2:3 population containing 94 lines.Then total 10,911 individuals from BC2F2、BC3F2 and BC4F2 were used to fine-map qRBSDV11,and then it was finally mapped to the region between two Indel markers HTA11-71 and HTA11-83,the physical distance was about 163 kb.There are 20 predicated ORFs in this region,and 3 ORFs with difference in the amino acid sequence between 9194 and SYN was classified as candidate genes.The fine mapping of qRBSDV11 established foundation to the map-based cloning of this gene.2.Genetic analysis and molecular mapping of QTL for small brown planthopper resistance in rice variety 9194.The RBSDVD-resistant variety 9194 displayed highly resistant to SBPH by modified seedbox screening test(MSST).To determine the mechanism underling SBPH resistance,we test the antixenosis and antibiosis of 9194 to SBPH.The results showed that the resistance against SBPH in 9194 might own to the antixenosis.To investigate whether the RBSDVD resistance of 9194 was due to its resistance against SBPH,we used 1049194/SYN F2:3 lines to identify QTLs for SBPH resistance.The frequency distribution of seedling mortality rate appeared continuously,with a range of 2.9-100%· It indicates multi-QTLs may control SBPH resistance in the population.Four QTLs,designated as qSBPH1,qSBPH5,qSBPH8 and qSBPH9,were detected on chromosomes 1,5,8 and 9,respectively.These QTLs explained 13.7%,11.0%,12.0%and 21.0%of the phenotypic variance with LOD scores of 2.72,2.78,2.15 and 2.85,respectively.The four QTLs jointly explained 57.7%of total phenotypic variance.Resistance alleles of the four QTLs were all derived from 9194.Among them,qSBPH9 was a major QTL.In our study,resistance to RBSDV disease in 9194 was obtained by resistance against both the virus and vector.Thus the QTLs conferring resistance to RBSDVD and SBPH in our study offer a new approach for breeding rice cultivars with RBSDVD-and SBPH-resistance through resistant genes pyramiding.