| Mollusc culture is a traditional marine aquacultute industry in China. It plays an important role in the economic development of the costal provinces. But since 1994, scallop aquaculture in China has been experiencing a large-scale mortality, and the culture industry suffered from a major great loss. The mortality was very serious and reached 80% or even more in many culture waters. The current large-scale mortality problem may result from the interaction of many different factors, such as foreign phathogens, environmental factors, and mollusc immunity. The etiological diversity of pathogens and the repeated appearance of new diseases led to develop new approaches for the control of diseases. Many researches have been carried out focused on the pathogens isolation and diagnosis, culture environment improvement, and culture technique optimization, and a lot of progress has been achieved.The cloning and expression of the genes related to disease-resistance are now considered to be a basic solution in the disease control because of their potential use in the development of therapeutic agents, study of immune defense mechanism and genetic improvement to increase the resistance to disease.In the present study, large scale EST sequencing method together with RACE technique was used to isolate and clone the genes involved in the immune defence from scallop cDNA libraries. Six full length cDNA sequences of immune-related genes were obtained, also another six partial cDNA sequences of immune–related genes were acquired.Three full length cDNA of heat shock protein 70 were cloned from three species of scallop, Chlamys farreri, Argopecten irradians and Mizuhopecten yessoensis. The molecular weight of the three HSP70s was ranged from 71.26kD to 71.80kD, and they were all acidic proteins. InterPro analysis revealed three classical signature motifs from these sequences. 3-D structure prediction showed that their N terminal corresponding the ATPase activity domain and the C terminal corresponding the substrate binding domain shared high similiarity with the heat shock protein 70 from human. Phylogenetic analysis of HSP 70 showed that all the molecules of scallop HSP 70 were clustered into one group. The relationships revealed by phylogenetic analysis of HSP 70 were consistant with the classical taxonomy. Semi-quantitive RT-PCR method was used to analyze the expression of heat shock protein 70 after the treatment of PAH and the challenge of gram negative bacteria. mRNA expression of heat shock protein 70 in scallop was up regulated significantly after stimulation of PAH, and there was a positive correlation between the mRNA expression and the concentration of PHA. While the challenge of gram negative bacteria could not regulate the expression of HSP 70.The full length cDNA of heat shock protein 90 was cloned from scallop, Mizuhopecten yessoensis. The molecular weight of the HSP90 was about 83.04kD, and it was also an acidic protein. The classical signature motif of HSP90 family was identified in the sequence of Mizuhopecten yessoensis HSP90 by InterPro analysis. 3-D structure prediction showed that both of the N terminal corresponding to the histidine kinase-like ATPase domain and the C terminal domain shared high similiarity with the 3-D structure of heat shock protein 90 from human. Semi-quantitive RT-PCR method was used to analyze the expression of heat shock protein 90 after the treatment of PAH and the challenge of gram negative bacteria. mRNA expression of heat shock protein 90 in scallop was up regulated significantly after stimulation of PAH, and there was a positive correlation between the mRNA expression and the concentration of PHA. While the challenge of gram negative bacteria could not regulate the expression of HSP 90. The full length or partial sequence of other immune-related genes, such as small heat shock protein, scavenger receptor (SR), cyclophilin A (CyPA), superoxide dismutase (SOD), metallothionein (MT), and peptidoglycan recognition protein (PGRP) were cloned from the sca... |
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