Bioinformatics Analysis ESTs Of Fenneropenaeus Chinensis And Meretrix Meretrix, Cloning And Expression Profiles Of Peroxinectin From Chinese Shrimp

Bioinformatics represents a new, growing area of intercrossing front discipline that uses computational and internet approaches to resolve biological questions. With the explosion of sequence and structural information available to researchers, the field of bioinformatics is playing an increasingly important role in the study of fundamental biological problems. In this dissertation, The author firstly constructed a marine bioinformatics platform based on Linux systems and cluster technology--Bioinformatics Linux Cluster (BLC), which includs Blast,Pfam,Phrap,Interpro etc.bio-software. Based on previous sequenced resoures, an Chinese Economical Marine Animals Genomics (CEMAG) web were constructed in intra-netsite. In order to screen more immune, growth and developmental related genes from Chinese shrimp and clam, different developmental phases cDNA libraries of clam were constructed. Subsequently, 2371, 1024, 1172 and 1060 expressed sequence tags (ESTs) were sequenced from haemolymph,eyestalk, ovary of shrimp and early juvenile larval of clam respectively. All these ESTs were analyzed, compared and annotated throughout constrcted BLC analytical platform. Further, 112 immune-related genes were discoveried from 971 unique genes of shrimp haemocytes ESTs including 5 main types according to the their functions: Antibacterial peptides (penaeidin, lysozyme, ALF, etc.), Pro-phenoloxidase system, (prophenoloxidase, serine proteinase, serine proteinase inhibitor, etc.), Clotting protein (agglutinin, hemagglutinin, transglutaminase, etc), Inter-cell signal communication (proexinectin, integrin), Chaperone proteins (HSP70, thioredoxin, thioredoxin peroxidase). And 150 genes involved in growth and development were also discoveried from 576 unique genes in clam larval ESTs, including structural genes, metabolize enzymes, immune-related genes and regulated genes etc. According to obtained bio-information, a 2370 bp full-length cDNA sequence of peroxinectin was cloned from F. chinensis by RACE method. The putative peptide sequence contains 684 amino acid residues. The predicted theoretical molecular mass and isoelectric point of deduced mature peptide is 74.58 KDa and 7.98 respectively. By motif scan, the putative peptide contain a peroxidase domain and a integrins bingding motif. This indicated peroxinectin of Chinese shrimp posses peroxidase activity and cell adhesion function. At the same time, this study develops relative quantitative real time PCR method to study through time, the mRNA expression profile of peroxinectin in haemocytes and hepatopancreas tissue of Chinese shrimp in response to no-activity bacteria infection. Research results showed constitutive expressional peroxinection mainly expressed in haemocytes tissue. The expression quantity changed rapidly during experimental times, from 6 hours to 72 hours. The change of peroxinectin is considered to be associated with increase cell adhesion activity and activation of proPO system. This indicated that peroxinectin is involved in exterior pathogen-defence mechanisms of Chinese shrimp.