Evolutionary Analysis Of The Main Grain Quality Traits In Indica Hybrid Rice Restorer Lines

The utilization of heterosis leads to a major breakthrough in rice yield,and contribute greatly to the food security in China.Over the past 50 years,remarkable progress has been made in rice yield during hybrid breeding.However,the research on genetic improvement of hybrid rice quality traits.Rice grain quality are complex traits,including processing quality,appearance quality,eating and cooking quality(ECQ)and nutritional quality.The main problem in hybrid rice grain quality is that the grain appearance quality and ECQ improved slowly during the hybrid breeding process.Rice grain appearance quality is mainly referring to grain shape and chalkiness,and there is a significant correlation between grain shape and chalkiness.The evaluation of rice ECQ is very hard,because factors such as amylose content,gel consistency,gelatinization temperature,starch viscosity,protein content and flavor play roles in determine rice ECQ.In recent years,consumers pay more attention to their health,thus the nutritional quality of rice has been concerned.Generally,the nutritional quality refers to rice protein content,lipid,micronutrient and the starch digestive characteristics(ie.resistant starch content).At present,there have been a lot of studies focus on the genetic regulation of rice quality traits,and a number of genes related to rice quality traits have been cloned,and the allelic variation of some key genes have been well studied.However,the allelic variation and utilization of these important genes in hybrid rice are not clear.Taking this as the starting point,this study collected 392 indica rice lines,including 58 two-line restorer lines(2-RL),218 three-line restorer lines(3-RL)and 116 inbred rice varieties(IRV),bred from 1970 to 2021,and systematically analyzed the main quality characters of these accessions.In order to understand the allelic variation and utilization of key genes related to grain quality traits within this germplasms,the whole genome re-sequencing data of these accessions were obtained and used for further analysis.Besides,genome-wide association analysis(GWAS)was carried out for rapid digestion starch and slow digestion starch parameters extracted from starch digestion characteristics.Also,the effect of major loci related to the above parameters was evaluated.The main conclusions of this study are as follows:1.The quality status of indica restorer lines were analyzed.For grain shape,there was a significant positive correlation between grain weight and grain length and width.The changes in grain quality from 1970 to 2021 of indica rice varied between different subgroups.For inbred rice varieties(IRV,116),the grain length increased gradually while grain width narrowed gradually,resulting in the high proportion of grain length-to-width ratio,which shows that the grain shape of rice become "thin and long" from "short and wide".Similarly,the grain length of the 2-restorer lines(2-RL,58)also increased gradually over time,but the increase range was smaller than that of the IRV accessions.The grain length and width of 3restorer lines(3-RL,218)did not change significantly over time.In terms of ECQ,there was a significant negative correlation between taste value and apparent amylose content(AAC)and setback value(SBV),while a significant positive correlation with breakdown viscosity(BDV).AAC was positively correlated with SBV and negatively correlated with BDV.There were no obvious trends in ECQ characters between 2-RL and 3-RL over time.However,for the IRV accessions,AAC showed a two stage change trend:in the first stage,from 1970 to 2000,the AAC decreased from?25%to?15%.Since 2000,the AAC maintained at about 15%.The gelatinization temperature(GT,Tp)of restorer line changed irregularly with the breeding process.The IRV showed clear change trends that most rice accessions bred in early stage have high Tp.From 2000,the number of low Tp accessions began to increase and maintained at a high level.There were no obvious change trends between rice subgroups and breeding yeas.2.The allelic variation and distribution patterns of key grain quality genes were analyzed by using whole gene re-sequencing data of 392 indica rice accessions.GWAS analysis showed that the main loci related to grain shape were mostly concentrated near GS3 and GW5 loci,indicating that these two genes play an important role in regulating grain shape within the accessions.The main loci related to ECQ are mostly concentrated near Wx and ALK genes,indicating that these two genes play key roles in regulating rice ECQ.The haplotypes of main grain shape genes(GS3,GW5,GS5 and GW6a)and chalkiness gene Chalk5,the distribution,as well as breeding years and regions were analyzed.The results showed that the haplotypes of GS3 and GW5 in the restorer lines changed irregularly over time,while GS5,GW6a and Chalk5 showed an obvious change trends.The proportion of slender grain control haplotype increased gradually,and the proportion of low chalkiness control haplotype also increased gradually.In IRVs,the haplotypes of GS3,GW5,GS5 and GW6a genes which control thin and long grains increased gradually over time.While the proportion of high chalkiness control haplotype of Chalk5 increased gradually.By using combination analysis,it was found that the grain shape showed two main types:one is "thin and long",the other is "short and wide".The functional alleles and haplotypes of Wx,ALK,SS?,SBE?b and Badh2 genes were analyzed.The results showed that the proportion of Wxb allele in IRVs and 2-RLs gradually increased with the breeding process,however,it did not change significantly in 3-RLs.The proportion of ALKb allele in IRVs increased gradually over time,but it did not change regularly in RLs.The haplotypes of SS? and SBE?b changed irregularly in all the accessions over time.The haplotype of Badh2 with aroma is mainly in IRVs,and its proportion increases gradually with the breeding process.3.GWAS was carried out for rapid digestion starch and slow digestion starch of 392 indica rice accessions.2 candidate segments were identified on chromosome 6,covering Wx and ALK genes respectively.An F2 segregation population was constructed by hybrid between a high resistant starch(RS)indica rice and a japonica rice.By using mmolecular marker screening and linkage analysis,10 loci related with RS were identified in this population.The LOD values of qRS60-2.2 and qRS60-6.2 QTLs were 7.99 and 9.21 respectively,and their contribution rates were 14.31%and 23.26%respectively.The qRS60-6.2 mapping interval is co-localization with the GWAS interval,which contains ALK gene.It was found that there was a significant positive correlation between RS content and Tp.In order to clarify the relationship between RS and ALK gene,the near isogenic line(NIL)NIL(ALKb)with low Tp and the NIL(ALKc)with high Tp were used for further analysis.The results showed that the RS content of rice grain from high-GT NIL was significantly higher than that of low-GT NIL rice.The above results show that ALK is not only the major gene controlling GT,but also contribute to rice RS.