| Zoysiagrass(Zoysia japonica)is a very important turf grass.It has been widely used in the construction of various lawn due to its strong stress resistance,extensive management and high ornamental properties..At present,many studies on Zoysiagrass including breeding of fine varieties,salt tolerance,chilling tolerance,disease prevention and development of molecular markers have been reported.The good quality of Zoysia is closely related to the microbial action in the rhizosphere environment.However,there are few studies on the salt tolerance and root microorganisms of this grass.Therefore,this research investigated the salt tolerance of Zoysia japonica from two aspects.The first part is to get genes information of fungi;the second part is to get numerous Zoysia salt tolerant genes.The main results are as follows:1.The transcriptome of rhizosphere and rhizosphere microorganisms grown in natural state was measured by using Illumina’s HiSeqTM2000 sequencing platform.A total of 68,024 unigenes were obtained after sequenced and de novo assembly.The average length was 802.07 bp,and the length of N50 was 1193 bp,5.49 Gb.By blast to the NR and Swiss-Prot databases,49,334 and 30,159 unigenes were annotated.In the total unigenes,28,284 unigenes of Zoysia japonica were isolated.Homology analysis showed that the relatives of Zoysia japonica were millet,sorghum,maize and rice.The results of gene function annotation revealed that 18,719 unigenes were enriched in 53 GO sub categories,and 10,300 unigenes were enriched in 24 COG categories,In the KEGG annotation,7,555 unigenes are enriched on the 124 metabolic pathways.In total unigenes,18,889 fungal sequences were isolated.In these fungi,the number of unigenes in 29 fungi exceeded 100,with the largest number of unigenes being Rhizophagus irregularis,and the two subspecies contained 4,074 unigenes.2.DAB staining analysis of Zoysia under salt stress treatment showed that the root surface was induced by hydrogen peroxide within 10 minutes;and then after 30 minutes later,the internal root is full of hydrogen peroxide,ROS related metabolic pathways have been activated.Zoysia ZjNHXl gene semi quantitative expression analysis showed that the expression of this gene was significantly up-regulated in 20 minutes,until 2 hours after treatment,the expression level was significantly higher than that before treatment.These results indicated that the metabolic pathways of SOS are also involved in the regulation of.20 minutes,ROS and SOS related metabolic pathways were activated,so sampling was taken at this time point.3.The control group(CK)and the salt stress treatment group(Case)were transcribed and sequenced respectively.After mixing and splicing,32,849 unigenes were obtained,with an average length of 1.107 BP and N50 of 1,781 bp.Based on the value analysis of RPKM,1,648 differentially expressed unigenes were found in two libraries.948 unigenes were up-regulated expressions and 700 unigenes were down-regulated.GO classification revealed that 1,455 differentially expressed genes were enriched in 73 subclasses,and 67 subclasses belonged to biochemical processes.The salt tolerance gene phylogenetics analysis showed that the salt tolerance of Zoysia japonica was closer to that of rice and millet.4.A total of 2,690 transcription factors were identified in unigenes based on sequence splicing,belonging to 80 transcription factor families.AP2/EREBP family had the most members,187 genes were obtained,followed by the bHLH family and the WRKY family,containing 158 and 137 genes,respectively.5.A NAC gene was cloned based on the transcriptome analysis of Zoysia japonica.Sequence analysis revealed that the gene sequence contained 1 complete open reading frames with a length of 1,332 BP,encoding a protein of 449 amino acids,containing a NAM domain.The real-time quantitative PCR result showed that the gene was induced by drought,high salinity and low temperature stress,and it might be an important gene in stress response. |
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