| Leymus mollis,a wild relative of wheat,is tolerant to salt and drought,and resistant to various diseases,which is considered as a useful genetic resource for wheat breeding. However,there is a lack of genetic information research in Leymus mollis. In order to understand genetic information of Leymus mollis and its molecular mechanisms of salt and drought tolerance,transcriptome sequencing of Leymus mollis leaf was performed by Illumina HiSeqTM 2000 high-throughput sequencing technology in this study. Functional annotation and classification of the unigenes were carried out based on the Clusters of Orthologous Groups(COG) and Gene Ontology(GO),and pathway annotation based on the Kyoto Encyclopedia of Genes and Genomes(KEGG) databases to further understand gene functions and regulation pathways. The results showed a total of 46 406 704 clean reads with Q20 of 96.96% and 55.29% of GC content were obtained from Leymus mollis leaf transcriptome library. These clean reads were assembled into 228 637 contigs with a N50 length of 438 bp. The contigs were further assembled into 112 846 unigenes with a N50 length of 1 319 bp. A total of 59 380(52.62%) unigenes were functional annotated successfully in the public protein databases. For species distribution,the majority of these aligned unigenes(40.69%) had first hits with sequences from barley,followed by Brachypodium distachyon and wheat. The results of COG analysis showed that a total of 15 786 COG annotated putative proteins were classified functionally into 25 categories. The cluster for “general function prediction only” represents the largest group among the 25 COG categories(7644,accounting for 48.42%),followed by “function unknown”(6444,accounting for 40.82%),“translation，ribosomal structure and biogenesis”(5689,accounting for 36.04%) and “transcription”(5467,accounting for 34.63%). GO analysis showed that 20 350 unigenes were classified into 44 functional groups that belong to biological process,cellular component and molecular function. For the hugest category of biological process,“metabolic process”(11 070,accounting for 54.40%) and “cellular process”(10 922,accounting for 53.67%) were the largest groups,followed by “single organism process”(6 153,accounting for 30.24%),“response to stimulus”(4 838,accounting for 23.77%),and “biological regulation”(3 399,accounting for 16.70%). Among the cellular component category,“cell”(14 589,accounting for 71.69%),“cell part”(14589,accounting for 71.69%),and “organelle”(12 630,accounting for 62.06%) were the most highly represented groups. In the category of molecular function,“binding”(10 230,accounting for 50.27%) and “catalitic activity”(10 024,accounting for 49.26%) were dominant. KEGG pathway analysis showed that 18 550 unigenes had significant matches in the database and were assigned to 128 pathways. The pathways most represented were “metabolism pathway”(5 168,accounting for 27.86%),“RNA transport”(3243,accounting for 17.48%),“mRNA surveillance pathway”(2864,accounting for 15.44%),“biosynthesis of secondary metabolites”(1 922,accounting for 10.36%) and “glycerophospholipid metabolic”(1 888,accounting for 10.18%). It is worth noting that unigenes involved in pathways of “plant-pathogen interaction” and “plant hormone signal transduction” were also abundant in this transcriptome datasets which included 1 367 and 861 unigenes,respectively. In addition,the key genes relative to transporting Na+ and genes responsive to oxidant stress were analyzed. The results showed that 15 unigenes functional annotated as Na+/H+ antiporter and 175 unigenes responsive to oxidant stress were found in the transcriptome database. Quantitative RT-PCR analysis was used to validate the expression level of the selected genes.These results will be helpful to further understand genetic information and molecular mechanisms of salt and drought tolerance of Leymus mollis. |
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