Map-based Cloning Of QTGW1.2b Regulating Grain Weight In Rice

Grain weight is a typical quantitative trait controlled by multiple genes with relatively small genetic effects.While major QTLs for grain weight have long been targeted for fine-mapping and cloning in rice,minor QTLs have not reveiced due attention.Identification of minor QTLs controlling TGW is a useful method to understand the genetic mechanism of seed weight in rice.In our previous studies,6NIL populations that derived from the indica rice Zhenshan 97 and Milyang 46 were developed for the dissection of QTLs in a 4.5 Mb region.Three minor QTLs for TGW were detected;including qTGW1.2a located in the 933.6 kb extending from RM11730 to RM11762,qTGW1.2b in the 418.8 kb extending from RM11781 to RM11800,and qTGW1.2c in the 2.1 Mb extending from RM11800 to RM11855.Among them,qTGW1.2a and qTGW1.2b mainly affect grain weight by grain length;qTGW1.2c mainly affects grain weight by grain width.This study aimed at qTGW1.2b,which QTL has the highest precision.New DNA markers were designed to increase the density of the target region,NIL populations with smaller segregating regions were developed to fine-map qTGW1.2b.Main results are summarized as the followings.According to the results of preliminary positioning of qTGW1.2b,a BC₂F₉ plant carrying a heterozygous segment interval RM212-RM11787 was selected from the Zhenshan 97/Milyang 46population.BC₂F₁₀ population was structured from the selfing progenies of the residual heterozygote.Using molecular markers to detect the genotype of BC₂F₁₀ population,two plants were screened,which carrying heterozygous segments covering the intervals RM11787 and Wn33304-RM11787,respectively.Two BC₂F₁₁1 populations were produced.Plants showing Zhenshan 97 homozygote and Milyang 46homozygote in the segregating regions were identified from the BC₂F₁₁1 population covering the interval RM11787.Selfing progenies of these homozygotes resulted in the production of NIL L2.L2 population was grown in two locations having distinct ecological conditions.In the mean time,the other BC₂F₁₀population was detected with molecular markers.Five plants covered the segments interval Wn33304-RM11787 were identified.Five sets of NILs named as L6,L7,L8,L9 and L10 were developed from the selfing progenies of the five plants.A QTL for TGW or GL was detected in each of the six populations.By comparing the segregating regions among the six NILs,qTGW1.2b was delimitated to a 371.5 kb region extending from RM11781 to Wn34526.L2 population detected significant QTL effects for grain weight and grain length with Milyang 46 allele increasing grain weight by 0.16 g,grain length by 0.026 mm,phenotypic variation explained was 15.41%and 11.74%.The effect of qTGW1.2b on grain was related to grain length.New Indel markers were developed to assay the genotype of cross-over regions of the L2 population.Newly identified homozygous segment was eliminated and qTGW1.2b was narrowed down to interval Wn34526–RM11787.Then,three plants carrying heterozygous segment covering this interval was selected from the BC₂F₁₃ population.Three BC₂F_14:154:15 NIL populations named as W1,W2 and W3 were established from the selfing plants showing Zhenshan 97 homozygote and Milyang 46 homozygote in the segregating regions.SAS procedure was used to analyze the phenotypic variation among Zhenshan97 genotypic group and Milyang 46 genotypic group.In each of the three NIL populations,significant variation among Zhenshan 97 genotypic group and Milyang 46 genotypic group was detected for TGW and GL.The MY46 allele increased grain weight by 0.12 g to 0.18 g,grain length by 0.021mm to 0.037mm.Thus qTGW1.2b was fine-mapped in a 44.0 kb region of Wn34323-Wn34367 by comparing the QTL interval of the three NILs.This region contains six open reading frames.By sequencing and expression analyses,it could be speculated that the target gene underlying qTGW1.2b was Os01g59410.