| As a self-pollinating plant soybean utilizated heterosis through the cytoplasmic male sterility / restoration(CMS / Rf)system.Parental outcrossing is mainly dependent on insects as the medium for pollen transmission.The outcrossing seed-setting rate of sterile lines plays an important role in soybean hybrids production,and it directly determines the yield or cost of the seed production.Therefore,it is of great significance to carry out research on the molecular mechanism of outcrossing rate and develope sterile lines with high outcrossing rate.In this study two soybean RN-type cytoplasmic male sterility(CMS)lines with significant differences in outcrossing rates were selected as the research objects.The mature buds at blossom period were used for transcriptome sequencing,and the data were analyzed by using bioinformatics methods.The genes with significant differences affecting the pollination efficiency of insects were verified by q RT-PCR(Quantitative real time polymerase chain reaction)in order to explore their affect on the outcrossing rate probably.The conclusions are as follow.1.Transcriptome sequencing analysis was performed on the flower buds of two soybean RN-type cytoplasmic male sterility lines with significant differences in outcrossing rate,and 1,925 differential genes were screened.Among them,1,361 differential genes were annotated to 30 branches of 3 major categories of Gene Ontology(GO),which was organisms’ processes,cellular components and molecular functions.Which mainly involves biological processes,catalytic activities,oxidoreductase activities,redox processes and carbohydrate metabolism processes.Meanwhile,864 differential genes were annotated into 114 metabolic pathways,which were classified by the Kyoto Encyclopedia of Genes and Genomes(KEGG),and mainly including metabolic pathway pathways,biosynthetic pathways of secondary metabolites,and plant hormone signal transduction pathways.2.Through the further analysis,13 biosynthetic pathways for secondary metabolites were revealed.By analyzing and identifying the differentially expressed genes of the flavonoid biosynthetic pathway which had significant differences and agreat number of enriched genes,it was found that the flavonol synthase(FLS)gene which affects flower color had a low expression level in high outcrossing rate sterile lines.However,it was in a high expression level in the low outcrossing rate sterile lines.So,it is speculated that the high expression level of FLS gene in sterile lines with low outcrossing rate could enhance the synthesis of flavonols and reduce the accumulation of anthocyanins.Thus,this could change the color of flowers and reduce the attraction to pollinating insects.3.35 differential genes which related to carbohydrates were screened,and 6genes with significant expression differences for verification were selected among them.The result showed that,the sucrose synthase(SUS11)gene closely related to nectar synthesis was found in a low expression level in the sterile lines with high outcrossing rate,and in a high expression level in the sterile lines with low outcrossing rate.So,it is speculated that when the SUS11 gene was at a low expression level,it can promote the synthesis of sugar in the nectar and increase the sugar content,which was conducive to attracting insect pollination and thus increasing the outcrossing rate of the sterile lines.It was discovered and identified the differential expression levels of the FLS and the SUS11 genes in sterile lines between high and low outcrossing rates,this result was helpful for the next step study on the relationship between biosynthetic pathways and outcrossing rates.Moreover,it expanded the thoughts for the further analysis of the molecular mechanism of the soybean outcrossing rate. |
