Detection Of Physiology And Biochemistry And Transcriptome Analysis In Senescence Leaves From Red Clover

Red clover is a leguminous forage with high nutritional and medicinal value.Its leaf senescence directly affects the yield and quality of red clover.At present,there are few studies on the senescence of red clover,and the regulatory genes and networks affecting the senescence of red clover leaves are still unclear.It is necessary to study the molecular mechanism of senescence of red clover leaves and find out the key regulatory genes of senescence of red clover leaves.The physiological and biochemical indexes of mature and senescent leaves of red clover were tested,including chlorophyll content,carotenoid content,soluble sugar content,proline content,soluble protein content and malondialdehyde content.The changes of physiological and biochemical indexes during senescence of red clover leaves were analyzed.The second generation sequencing technology was used to sequence and analyze the transcriptome of mature and senescent leaves of red clover.The differentially expressed genes in mature and senescent leaves of red clover were screened.The changes of gene expression and metabolic pathway during senescence of red clover leaves were analyzed.It is helpful to reveal the main pathway of senescence mechanism of red clover leaves and provide a basis for exploring its senescence mechanism.The theoretical basis will play a guiding role in tapping potential genes for delaying leaf senescence of red clover.The main results are as follows:1.Compared with mature leaves,chlorophyll content,carotenoid content and soluble protein content of senile red clover leaves decreased significantly,while soluble sugar content,proline content and malondialdehyde content increased significantly.2.Through transcriptome sequencing analysis,314,043,664 raw reads were obtained,Q20 was 96.03-96.30%,Q30 was 90.28-90.81%,GC content was 40.63-41.17%.After filtering low-quality reads,299,574,840 clean reads were obtained.3.By comparing and splicing the reference genome of red clover with that of red clover,35,067 genes were detected in the two developmental stages of red clover.Through differential expression analysis,481 differentially expressed genes were screened,including 95 up-regulated genes and 386 down-regulated genes.Twenty genes were randomly selected for qRT-PCR detection.The results showed that the expression trend of these genes was consistent with the results of transcriptome sequencing analysis,which indicated the reliability of transcriptome sequencing for leaf senescence analysis of red clover.4.The classification of GO functional annotations showed that differentially expressed genes were enriched into three categories and 30 subcategories.KEGG annotation indicated that the differentially expressed genes were mainly concentrated in biosynthesis of secondary metabolites,cyanoamino acid metabolism,plant hormone signal transduction and phenylpropane biosynthesis,suggesting that these processes may play an important role in the senescence of red clover leaves.