EST Analysis Of Argopecten Irradians And Chlamys Farreri And Cloning And Expression Of C-type Lectin Gene In Chlamys Farreri

There is a big industry of scallop culture in China, and Bay Scallop (Argopecten irradians) and Zhikong Scallop (Chlamys farreri) are the most important cultured marine mollusks. In the past years, scallop culture in China has been experiencing from a mortality problem, and suffering from a great economic loss. Although it is still unclear about the causes for the scallop mortality, deterioration water quality and deterioration of the stock are suspected to be the most important causes. With the improvement and control of the marine environment, enhancing the disease resistance of scallop is commonly believed to be the resolution to the control of the disease. The cloning and expression of the genes involved in immune defence 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. Expressed Sequence Tag (EST) is an efficient method for high throughput gene screening and identifying, which was applied widely in humans and other species. In the present study, cDNA libraries from C. farreri and A. irradians were constructed and EST analysis was performed in an effort to establish and provide a well-characterized EST resource for the new gene identification and their expression patterns. mRNA was isolated from the whole tissue of both healthy Bay Scallop and Zhikong Scallop stimulated by Vibrio anguillarum, and then was reverse-transcripted into cDNA. The double strands cDNA were ligated into a vector to construct the λ phagemid cDNA libraries. Two cDNA libraries were - 3 -胥 炜 海湾扇贝和栉孔扇贝EST分析及C型凝集素基因的克隆与分析 硕士学位论文 obtained with the average inserts about 1.2kb. 7680 positive clones were selected from each cDNA library and then were sequenced from the 5' ends. After proceed with a serial bioinformatics methods including vector masking, files format translating, rejecting the sequence shorter than 100bp and low qualities, 4991 and 5123 ESTs were obtained from Bay Scallop and Zhikong Scallop cDNA library respectively. The average sequencing length of Bay Scallop ESTs and Zhikong Scallop ESTs were 462.01bp and 392.52bp respectively, while the actual length of sequencing fragments were 347.62bp and 360.06bp respectively. Clustered with Phrap program, 637 contigs and 2138 singletons were obtained from Bay Scallop ESTs, while 584 contigs and 2762 singletons from Zhikong Scallop. Blast analysis showed that in Bay Scallop ESTs, 259 contigs and 585 singletons have a high homology with some genes in GenBank (E-value<0.005) while in Zhikong Scallop ESTs, 332 contigs and 725 singletons can find their homologous genes in GenBank. But most of them could not be matched with any known genes, 56% in both Bay Scallop ESTs and Zhikong Scallop ESTs. About 1% sequences of both Bay Scallop data and Zhikong Scallop data have high homology with cell division related genes; 2% Bay Scallop ESTs and 3% Zhikong Scallop ESTs were homologous with cell signal and communication; 9% Bay Scllop ESTs and 8% in Zhikong Scallop ESTs were highly homologous with cell structure and motility; 12% Bay Scllop ESTs and 5% Zhikong Scallop ESTs have homology with gene and protein expression; 7% Bay Scllop ESTs and 8% Zhikong Scallop ESTs were found to be homologous with metabolism; another 7% Bay Scllop ESTs and 6% Zhikong Scallop ESTs have some homology with the genes which have no clear function. The most important part of these data is the ESTs homologous with cell and organism defence, which were 6% of Bay Scallop ESTs and 13% of Zhikong Scallop ESTs. Some important immunity related genes were found in the data including defensins, lysozymes, lectins, heat shock proteins, peroxidases, serine proteinases and metalloproteases and so on. This result provides...