| Litopeneaus vannamei (L.vannamei), also known as pacific white shrimp, white prawnand whiteleg shrimp, has the advantages of rapid growth rate, strong immune defenses,intensive farming adaption and high nutrition that make it be the mainly cultivated shrimpspecies throughout the world. However, in our country the culture of L.vannamei is stillrestricted by nutrition, breed and disease. To study the fundamental mechanism of shrimpmuscle growth and development and immune system could provide theory basis forimproving growth rate and enhancing resistance of L.vannamei, that has significant role inefficient and healthy shrimp aquaculture.In study, homologous cloning was employed to attained partial cDNA sequence of p38inL.vannamei and bioinformatic analysis was performed on it. Besides, the expression of p38indifferent tissues and molt stages was detected by real-time quantitative PCR, which was alsoapplied to measure the expression of genes related muscle after silencing the expression ofp38by dsRNAi and inhibiting the enzyme activity of p38by inhibitor in vivo. At last, theexpression patterns of p38after bacterial and virus infection and the change of viral copies inL.vannamei after infected with WSSV were analyzed.According to the results, partial cDNA sequence of p38was acquired, which has1206bpcontained1098bp open reading frame that encodes365amino acids with molecular weight41.87kDa, pI6.09and no signal peptide. p38is highly conserved with others, which includesS_TKc domain and TGY motif in activation loop. It also contains11subdomains which hassignature sequences of MAPK family and p38subfamily. Tissue distribution analysis foundthat p38highly expressed in hepatopancreas and muscle, which indicated it would play animportant role in immune response and muscle growth and development of L.vannamei.Though analyzing the expression pattern of p38during molt stages of L.vannamei, it wasfound that p38had high expression in intermolt on mRNA and protein levels. Meanwhile, thecontent of phosphorylated p38in intermolt was obvious higher than other stages. Theseresults suggested that p38may involve in muscle growth and development at the stage ofintermolt. Additionally, the silencing of p38expression had significant effects ontranscriptional expression of muslce related genes (MHC, MLC, TPMS, TPO and α-actin),while inhibition of p38only effected the expression of MLC and TPO and had no significanteffect on expression of MHC, TPMS and α-actin. These implied that p38could participate in muscle growth and development through regulating the expression of muscle related genes byvarious ways.What is more, to explore the functions of p38in immune response, we detected theexpression of p38in hepatopancreas after L.vannamei infected by different pathogens. Theinfection of WSSV had no significance effect on expression of p38except at24thhour afterinfected. In addition, the expression of p38showed an obvious decline in12hours beforesignificant increase at24thhour after injection of Vibrio Parahaemolyticus, while it hadsignificant increase after infection by Micrococcus Lysoleikticus and Vibrio Anguillarum.Therefore, p38could play a vital role in shrimp against bacterial infection, whereas its role indefensing viral infection needs to be further research. Lastly, the copies of WSSV inL.vannamei after infection was be detected. We found that it remained at certain number in24hours after infection, followed by a drastic increase, which indicated that viruses replicatemainly at36-48hours after virus infection.As the results described above, we cloned partial cDNA sequence of p38and found itinvolved in shrimp muscle growth and development by various ways and played an importantrole in defensing against bacterial infection in L.vannamei. This thesis explored the functionsof p38in L.vannamei muscle growth and development and immune response, which providestheory basis for throughout and systematic researches in muscle growth and development andimmune system on L.vannamei. |
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