Fine Mapping Of The Flag Leaf Angle QFla-8-2 Locus In Rice(Oryza Sativa L.)

Recently,the area planted with hybrid rice occupied 60%of the total area of rice in China,150,000 hm2 of hybrid seeds should be produced each year.However,the low outcrossing rate of sterile lines and high cost of seed production are important factors for restricting the development of hybrid rice.In order to improve outcrossing rate,it is necessary to spray GA3 and cut partial flag leaf in the process of seed production.Cutting the flag leaf not only requires a large amount of workers but also high-technique operation.Becides,mechanization development of the hybrid rice seed production is also seriously hindered by a lot of manual operation.By cultivating male sterile lines and maintainer lines of the large flag leaf angle to improve the growth profile of flag leaf in rice,the female stigma is becoming more easer to accept male pollen.It is very important to solve the problems of artificial leaf cutting and low outcrossing rate,and finally accelerate the mechanization process of hybrid rice seed production.To better understand the genetic mechanism of rice large flag leaf angle,we fine mapped the flag leaf angle QTL qFla-8 detected by BC1F1 population derived from Japonica rice maintainer line 863B and large flag leaf angle germplasm A7444,and finally predicted the candidate gene of target chromosome region.The initial material was BC3F3 population came from 863B/A7444//863B.Firstly,heterozygous lines at qFla-8 locus were identified in BC3F3 population.We further narrowed down the target region by using 9 SSR markers and 172 plants.Than 68 heterozygous lines at qFla-8-2 locus were identified and we harvested the selfed seed to enlarge the separation population.Secondly,the target gene was limited within 100 kb chromosome segment by using more newly added molecular markers.Thirdly,some candidate genes were predicted in the target segment through the NCBI database.The main results were as follows:1.There were two mutex linked loci qFla-8-1 and qFla-8-2 in the initial QTL region.Among them,qFla-8-1 was located between the SSR markers RM6215 and RM3153,explaining 22.33%phenotypic variation.Positive allele existed in 863B and had 8.59 LOD value.qFla-8-2 was located between the SSR markers RM1309 and RM3153,explaining 23.81%phenotypic variation.Positive allele came from A7444 and had 8.83 LOD value.Physical distance of qFla-8-1 is 76 kb and the other is 460 kb?2.Genetic analysis showed that the large flag leaf angle was recessive with the small flag leaf angle.438 individuals with great large flag leaf angle were selected from BC3F4 population to screene recombinants by using flanked markers RM1309 and RM3491.We finally got 21 recombinants at both sides.There were 13 recombinants between RM1309 and qFla-8-2,8 recombinants between RM3491 and gFla-8-2.In order to obtain more markers,35 InDel primers and 12 SSR primers were designed,of which only two InDel and two SSR primers had polymorphism.Using the two InDel markers and two SSR markers to analyze 21 recombinants,it indicated that there were 12 recombinants between RM23065 and qFla-8-2,9 recombinants between Z5 and qFla-8-2,5 recombinants between Z7 and qFla-8-2,4 recombinants between RM23071 and qFla-8-2.Therefore,the gene qFla-8-2 was mapped to about 67 kb segment between InDel marker Z7 and SSR marker RM23071.3.This 67 kb region contains three predicted genes based on NCBI:Os08g0408200 has ten exons,encoding WD40 domain containing protein similar to GAMYB-binding protein;Os08g0408300 has one exon,encoding hypothetical protein;Os08g0408500 has one exon,encoding APETALA2-like protein.Os08g0408200 was considered as the most possible candidate gene,further transgenic experiment or population genetics experiment is ongoing.