| Ovomermis sinensis is an important natural enemy of insects and has a great potential in biological controls of pests in the agricultures, vegetables and forest. As the Mermithidae of insect parasite, it has a specific sexual decision mechanism, which is determined by quantity of the nutrient through the parasitic stage. As environmental pollution, human destruction, and the limited resources of Mermithidae, these of bio-control resources nearly disappeared. Therefore, many scientists tried to culture the Mermithidae in vitro in order to solve the problem. However, this method is not successful, the reason is that the Mermithidae cultivated in vitro is not maturation in sex. We have known a little about the molecular mechanism of sex-determination about Mermithidae by now. On the other hand, Mermithidae is a valuable bio-control resource. Thus it is important for us to research the sexual decision mechanism of O. sinensis. For one thing, this research has the theory significance. For another thing, in order to carry out the culture in vitro of O. sinensis and provide insect enemy resources for the agricultural production, this study has the practice significance.The DEAD-box family genes vasa and p68 have been studied in many species from yeast to human. As the germ cell marker, vasa always use for the research of reproduction such as the production and removal of primordial germ cell and so on. The production signals of p68 mRNA are always detected in different tissues among distinct species. DEAD-box family proteins are the ATP dependent RNA helecases that contain four sub-families; they all involve in many aspects of the reproduction, developments, organ differentiation and cell proliferation. Predecessors in our lab have cloned laf-1 gene in O. sinensis and detected the expression of laf-1 mRNA through the all developmental periods. The results revealed that the signals were much higher during the sex determination period compared with other periods. In addition, the laf-1 mRNA has been detected in different tissues, the consequence manifested that the expression signals were appeared in many tissues. We have cloned Tra-1 which has essential function for the gender development in female adults.We have not understood whether the other genes of DEAD-box family exist and the roles of these genes in O. sinensis. In addition, O. sinensis has complex sex differentiation molecular mechanism. In order to research the mechanism of Mermithidae, we need more support of gene research. Therefore, to understand the sexual determination and reproduction of Mermithidae, we studied this special mechanism in molecular field as followings.1. Through the methods of RACE and RT-PCR, we have cloned the vasa of O. sinensis, which are 3053 base pairs for the full length. Its ORF consists 2439bp, and it encodes 813 amino acids. The results by the RT-PCR through the specific primers showed that vasa mRNA was expressed specifically in the gonads of both testis and ovary. However, in other tissues such as head, tail and body wall, there is no vasa mRNA expression signals.2. We have detected the expression of vasa mRNA through the defferent developmental stages of O. sinensis by Real Time PCR. The outcome showed vasa mRNA expression was high in embryo period, inceased firstly during the proliferation of testis and ovary, and was the highest during the fifth day of extrusion from the host(P< 0.05). It was decreased as testis and ovary matured. After leaving the host, the trends of the transcription signals showed that rise firstly, and then decline, so we speculated that vasa had the relationship with gonads development. The analysis results of early embyo development in O. sinensis showed that vasa mRNA was expressed during the all early embyo stages, and the relative expression increased firstly and decreased finally. It suggests the vasa of O. sinensis is a maternal gene and plays an essential role during early embryo development.3. The full length of p68 (Ddx5) of O. sinensis which is 2123bp was obtained by the method of SMART RACE. The putative amino acid contained the domains of DEXDc and HELICc.4. Through the method of RT-PCR, we have detected the expression of p68 mRNA in different tissues such as head, tail, testis, ovary and body wall in O. sinensis. Earlier reports showed that P68 was expressed highly in mice male germcells. The results showed that p68 gene was expressed in all these parts, we speculated that the activity of RNA helicase has functioned widely.
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