| There are two chapters in this thesis. In chapter one there is an elementary study on the sensitivity of Bombyx mori larvae to Dendrolimus punctatus cypovirus strain Hunan (DpCPV-HN). DpCPV-HN was cloned in vivo and identified as a type 1 cypovirus. The fourth or fifth - day - post-hatching F1 and F2 silkworms from generation F1 that had previously been infected by DpCPV-HN, and two other type 1 cypoviruses from B. mori ( BmCPV-1 ) and D. punctatus ( DpCPV-HN ) were selected for the infection test. The results indicated that the breed of B. mori tested in this study could be infected by DpCPV-HN, of which the virulence to silkworms was inferior to that of BmCPV-1, and the difference of virulence between the two CPVs was distinct. Based on the accumulative larval mortalities of F1 and F2 B. mori infected by DpCPV-HN over 28 days post - infection, the 50% lethal doses (LD50) of DpCPV-HN for F] and F2 generations were 885 and 18 CPBs, respectively, which indicated that F2 silkworms were more sensitive to DpCPV-HN than those of FI. To some degree, the ratio of cocoons, ratio of pupae, ratio of moths to pupae, cocoon weight, cocoon shell weight, ratio of cocoon shell weight, and number of eggs from single female moths of the F1 and F2 generations declined after infection by DpCPV-HN. The correlations between ratio of moths to pupae of F2, F1 B. mori and inoculation doses of DpCPV-HN were significantly and highly significantly negatively relative, respectively. The correlations between cocoon weight, cocoon shell weight, ratio of cocoon shell weight, number of eggs from single female moths of silkworms and inoculation doses of DpCPV-HN were not obvious.In chapter two, the nucleotide sequences of genome segments 6, 7 and 10 of DpCPV-HN were amplified by RT-PCR, and the cDNAs were cloned in E. coliDH5a, determined and analysed, respectively, and the cDNA encoding p50C259 of DpCPV-HN was then expressed in E. coli BL21. Gene S10 was 944 nucleotides in length and encoded a polyhedrin of 248 amino acids (28.5 kD), and the complementary sequences 5'- AGTAA-3' and 5'- TTACT- 3' in the polyhedrin gene plus strand were found. When compared with BmCPV-1 S10, DpCPV-HN S10 had 91.2% identity in the nucleotide level and 98.0% identity in the amino acid level. When the nucleotide sequence of DpCPV-HN S10 was compared with that of DpCPV-JX polyhedrin gene ORF, five transition substitutions at nucleotides 122, 563, 667, 734 and 740 and one transversion substitution at nucleotide 575 were found, but only the transition mutation at nucleotide 667 resulted in a replacement of Lys209 in DpCPV-JX polyhedrin by Arg209 in DpCPV-HN polyhedrin. Gene S7, 1501 nucleotides, encoded a protein of 448 amino acids (49.8 kD; p50), and one nuclear localization signal ( NLS ) was found in p50. Comparison of the amino acid sequence of p50 with GenBank and EMBL databases revealed some similarity to DnaK-like protein of Mycoplasma hominis. When compared with BmCPV-1 S7, DpCPV-HN S7 had 87.0% identity in the nucleotide level and 92.8% identity in the amino acid level. The ORF of gene S6 was 1686 nucleotides in length and encoded a protein of 561 amino acids (63.7 kD; p64). The amino acid sequence of p64, had a high leucine content (10%), and contained a leucine zipper motif and one ATP/GTP-binding site motif and two NLS, suggesting that p64 may possess nucleic-acid-binding activity and might play an important role in CPV replication. Comparison of the amino acid sequence of p64 with GenBank and EMBL databases revealed some similarity to Rice ragged stunt virus ( RRSV ) Pns7 encoded by RRSV S7. When compared with BmCPV-1 S6, DpCPV-HN S6 had 83.7% identity in the nucleotide level and 92.5% identity in the amino acid level. The cDNA encodingamino acid residues 190 to 448 of p50 (p50C259) from DpCPV-HN was cloned, and then inserted into pET-28a expressive vector after being digested with BamH I and Hind III to construct pET-28a/S7(591-1371) recombinant expression vector. Expression of the...
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