Screening And Identification Of The Differentially Expressed Genes Between Different Developmental Stages And Analysis Of New Genes In Schistosoma Japonicum

Schistosomiasis, caused by schistosome, is the wide spread zoonosis that seriously threatens the human and animal health. Schistosome exhibites complicated characteristic growth and development. During its life cycle, schistosome presents dramatic changes in its biology and morphology. It is believed that these biological and morphological changes are accompanied by differential gene expression pattern at the various stages, which result in special metabolism and development. Based on the above features, the screening and identification of differentially expressed genes of Schistosoma japonicum were carried out, and several differentially expressed new genes were cloned and analyzed. Our results are useful for better understanding schistosome special growth mechanism and interaction with host on molecular level, and to open a new path for preventing and curing schistosomiasis.1. Screening and identification of Schistosoma japonicum differentially expressed genes between different developmental stages.Previously undiscovered differentially expressed genes cDNA libraries from Sj cercariae, egg and adult worm were successfully constructed by suppressed subtractive hybridization technique. The evaluation results indicated that the three subtracted cDNA library were high quality, and were applicable to isolate Sj differentially expressed genes on the whole genome scale. The 257 ESTs of differentially expressed genes from three subtracted cDNA libraries were analyzed. The result suggested that genes from cercariae subtracted cDNA library mainly involved in movement, energy metabolism, transcription regulation, and pathogenesis; genes from egg subtracted cDNA library largely involed in signal transduction, cell adhesion, protein and carbohydrate metabolism, and response to oxidative stress; and genes from adult worm subtracted cDNA library mostly involved in protein biosynthesis , transport, proteolysis, and worm movementThe cDNA microarray hybridization was used to screen and identify differentially expressed genes. A cDNA microarray was fabricated with 3111 gene clones selected from cercariae, egg and adult worm subtracted cDNA library. Microarray hybridization results indicated that 1620 gene clones presented differential expression between tester and control, including 374 cercariae differentially expressed genes, 701 egg differentially expressed genes and 545 adult worm differentially expressed genes. Several nonredundent differential expression gene clones were verified by RT-PCR and Northern blot, which demonstrated that these clones were indeed differentially expressed, and confirmed the cDNA microarray results, thereby supporting the reliability of the system. Our results indicated that it is feasible that SSH combined with cDNA microarray to isolate and identify differentially expressed genes. A total of 108 clones from above differentially expressed genes were sequenced. The results of sequence analysis indicated that these clones represented 44 unigenes. Among these unigenes, 5 schistosome known genes have been previously identified as differentially expressed genes, and 6 schistosome known genes were identified as differentially expressed genes in our study. Furthermore, in the remaining 33 unigenes, one is similar to a hypothetical protein gene, one is homologous to...