High-Throughput Transcriptome Analysis And Immune Gene Discovery Of Macrobrachium Nipponense

The freshwater prawn,Macrobrachium nipponense,is an economically important species cultured in China.Due to the short breeding period and high market prices,Macrobrachium nipponense is populared with the aquatic breeding farmers.But in recent years, the frequent occurrence of prawn diseases caused by bacteria has led to serious economic loss of prawn farming.So the study of Macrobrachium nipponense immune defense mechanism on the molecular level,is very important for the disease prevention and control.In this study, using Illumina high-throughput sequencing technology,Successfully constructed a transcriptome library,and four pair DGE libraries of different bacterial stimulation time and different organization from Macrobrachium nipponense.The total transcriptome library was assembled from eight transcriptome libraries.By using blastx algorithm,the functional annotation and metabolic pathway analysis of Macrobrachium nipponense total transcriptome Library was completed in the public database(COG,GO,KEGG,KOG,Pfam,Swissprot and nr).Subsequently, some important immune genes were screened.In order to understand the temporal and spatial specificity expression of immune genes in Macrobrachium nipponense,we did the classification comparison among the 4 pairs DGE libraries in different organs and different stimulated time.This work is contribute to reveal the immune mechanism of Macrobrachium nipponense from the molecular biology level.In the process of transcriptional analysis, we identified the transcription factors Litaf and TNF, and analyzed the two transcription factors in order to verify whether they are related to immunity. The main results as follows:1.Macrobrachium nipponense transcriptome library was successfully constructed,which contain 142560 unigenes. A total of 43038 genes were annotated, accounting for 30.20% of the total unigenes of the total group.1240 of these genes were associated with the immune signaling pathway.Through the analysis of immune related genes, identified the main pattern recognition receptors, signal transduction factors and immune effect factors exist in invertebrates.But d FADD and Dredd gene were not identified in Macrobrachium nipponense.According to the screening results of immune genes, preliminary drew the Macrobrachium nipponense immune pathway.2.We compared the transcriptome between different stimulation time and different tissues first and DEG analysis of immune related genes afterwards.The specific expression of immune genes between the haemolymph and hepatopancreas was identified.The specific expression of immune genes after bacterial stimulation was identified too.The temporal and spatial specificity expression of immune genes in Macrobrachium nipponense was preliminary cleared.3.This is the first study in shrimp to identify TNF super family member Mn TNF.The quantitative analysis results show that the highest expression level in gill;Quantitative analysis of Mn TNF upstream potential transcription factor Mn Litaf.The results showed that the high expression in hemolymph.After bacterial stimulation, the expression of TNF and Litaf were significantly up-regulated,suggesting that they play an important role in Macrobrachium nipponense immunity.In summary,this research enrich the genetic database of Macrobrachium nipponense, provides a theoretical basis for Macrobrachium nipponense of disease prevention and healthy aquaculture.