Analysis The Difference Of Spermary Gene Expression Of The Diamondback Moth Parasitized By Different Parasitoid

Cotesia plutellae(Kurdjumov)(Hymenoptera : Braconidae)and Diadegma semiclausum Hellen (Hymenoptera: Ichneumonidae)are the major larval parasitoid of the diamondback moth, Plutella xylostella (Linnaeus) (Lepidoptera: Plutellidae), an important insect pest of crucifer vegetable crops. In order to provide theoretical evidences in better understanding of the expression and physiological effects of the two species polydnavirus, Bracovirus and Ichnovirus , in the same host, and the physiological regulation mechanism of the Bracovirus and Ichnovirus on the same host in the two system of C. plutellae—P. xylostella and D. semiclausum-- P. xylostella, our study was investigated by analyzing the difference of spermary genes expression, isolating and cloning the related genes that arrest the development of of the diamondback moth by differential display method which developed recently.Total RNAs were extracted from spermary of normal diamondback moth, the diamondback moth parasitized by C. plutellae and the diamondback moth parasitized by D. semiclausum, separately. cDNA fragments were amplified by PCR with arbitrary primers 1-8, and anchor primers T₁₁M (M is G/A/G) by differential display PCR. The amplified PCR products were separated on 1% gelose gel electrophoresis. The special cDNA bands for polydnavirus were recovered from the gelose gel, and were reamplified using the corresponding primer sets, sub-cloned, sequenced, and confirmed. These results suggest that more than ten cDNA bands, which might represent the polydnavirus differentially expressed genes, were obtained by DD-PCR method. Seven of them were selected to reamplification by PCR, and were cloned into the PT18-T plasmids. The positive clones were identified by screening of blue-white dots. The cloned cDNA fragments were sequenced When compared arose partial nucleotide sequences with the sequetxxs deposited in the GenBank data, one cDNA fragment showed 100% sequence homology to the Oryza sativa (japonica cultivar-group) gene, suggested that it was contaminated, and the rest cDNA fragments all did not show any significant homology. These cDNA fragments confirmed by reamplified using the corresponding primer sets, in three kinds spermary all have fragments can be amplified.These results suggest that more then ten cDNA fragments, which might be related polydnavirus differentially expressed genes, are identified by the method of DD-PCR. But none of those fragments was confirmed just is the related gene of polydnavirus yet.