Analyses Of Leaf Transcriptome And SRNAs Under Heat Stress In Narcissus

Narcissus tazetta var.chinensis is one of the ten traditional famous flowers with high ornamental value in China and is also the provincial flower of Fujian,it can be cultivated under warm and wet climate,too high temperature can result in shortenning of flowering period and blooming time.In recent decades,the greenhouse effect has resulted in rising of global temperature year after year,and become one of important characteristics of future climate,Zhangzhou City is one of main production regions of Narcissus,transcriptome and small RNAs of leaves of plant of Narcissus treated under normal condition?at 15 °C?and high temperature stress conditions?at 30 °C and 35 °C?were sequened and analysized by BGISEQ-500 sequencing for exploring pathways of response to heat stress and mechanism of heat-tolerance of Narcissus,which will provide scientific evidences for finding heat-tolerant genes and developing heat-resistant cultivars of Narcissus.The results were as follows:1 Transcriptome analysis of leaves of plants of Narcissus treated under high temperature stressesTotal RNAs?3 replicates?of Narcissus leaves of plants treated under normal condition?at 15 °C,as control,and high temperature?at 30 °C and 35 °C?conditions?were extracted to construct their respondingcDNA libraries for BGISEQ-500 sequencing.The results of transcriptome sequencing showed that 101224,102603 and 103049 transcripts were obtained in the three resulted cDNA libraries,respectively,and that112160 unigenes whose total length was 128733815 bp,whose average length was 1147 bp,whose N50 and GC content were 1770 bp and42.33% were obtained by emerging of cDNA libraries,and that 19022 loci of SSR and 71619 CDS sequences were found.and that 27007 differentially expressed genes?DEGs?including 6029 up-regulated genes and 6366 down-regulated genes in T15 and T30 combination,and 10788up-regulated genes and 12411 down-regulated genes in T15 and T35 combination,and 3090 up-regulated genes and 4719 down-regulated genes in T30 and T35 combination were found in three temperature treatments.The results of GO enrichment of DEGs showed that photosynthesis system and membrane system of leaves of Narcissus plants treated under heat temperature stress were significantly affected.The results of KEGG showed that there were the largest number of differential genes in metabolic pathways,that compared with control combination,the two pathways with significant enrichment under high temperature treatment include secondary biosynthesis,photosynthesis,glycosaminoglycan degradation,metabolic pathways?ko01100?,glyoxylic acid and dicarboxylic acid.Eight pathways including metabolism,phenylpropanoid biosynthesis,photosynthesis-antennaprotein,and monoterpenoid biosynthesis.Quantitative results show that the sequencing results have high accuracy,and Hsf,HSP and TPS genes play an important role in the heat stress response of Narcissus.2 Small RNA analysis of high temperature responses in NarcissusSmall RNA sequencing using the BGISEQ-500 sequencing platform of the expriment material,which same as the used for transcriptome sequencing.The results of sRNAs sequencing showed that total of 96 known miRNAs and 25 novel miRNAs and 52286 predicted siRNAs were obtained in three libraries.Analysis of differentially expressed?DE?siRNAs indicated that some siRNAs play an important role for high temperature treated responses of Narcissus,e.g.,in the predicted siRNAs,the types and numbers of predicted siRNAs gradually increases as the treatment temperature increases.Analysis of DE miRNAs suggestings that some miRNA families were specific for different high temperature treated responses,and different members of the same family expressed different expression patterns under different degrees of heat stress,such as novel_mir7,miR166 u,miR172a₂,which may be play an important role in the heat stress process.The GO and KEGG pathway enrichment analysis of DE miRNAs targets showed that the number of the plant hormone signal transduction pathway was the highest in the three comparison groups,and mRNA surveillance pathway,RNA polymerase,the RNA degradation was significantly enriched as well,but thephingolipid metabolism pathway occurred in the T15_vs_T30 group,indicating that it may play an important role in the response of Narcissus to high temperature stress.Sum as there may be multiple response pathways while under high temperature stress in Narcissus,especially phytohormone signaling pathway and reactive oxygen species pathway.The results of qRT-PCR verification showed that the accuracy of miRNAs sequencing data was high,and that miR160,miR168 and miR167 may play a role in heat stress of Narcissus.