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Cloning and Characterization of BmPTERP and Bm3HCDH Genes of Silkworm, Bombyx Mori

Author: WangXiu
Tutor: GuoXiJie
School: Jiangsu University of Science and Technology
Course: Biochemistry and Molecular Biology
Keywords: Bombyx mori cytoplasmic polyhedrosis virus rapid amplification of cDNA ends real-time quantitative PCR
CLC: Q78
Type: Master's thesis
Year: 2011
Downloads: 43
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Abstract


Bombyx mori cytoplasmic polyhedrosis virus (BmCPV), one of the important viral pathogens for the silkworm disease, causes harm to product of silkworm and enormous damages to the sericultural industry. Studies on genes related to silkworm infected with BmCPV and understanding of silkworm molecular mechanism of BmCPV infection is crucial to the prevention of BmCPV-caused silkworm disease and the development of new medicine of silkworm.In our previous work, large number of differentially expressed genes were identified in the midgut of infected BmCPV by using microarry analysis. Based on the EST information of this microarry analysis, the cDNA of two genes were cloned using the strategy of rapid amplification of cDNA ends. Concurrently bioinformatic methods were applied to analyze the obtained sequences of these genes. The conclusion are as follows:1. Molecular cloning and characterization analysis of phosphotriesterase-related protein gene of silkworm,Bombyx mori (BmPTERP).BmPTERP consists of 1349 bp and contains a 131 bp 5’UTR and a 168-bp 3’UTR. The open reading frame which locates in the region from 132 bp to 1181 bp of cDNA encodes a 350 amino acid protein. The molecular weight of the putative BmPTERP was 39.03-KDa and isoelectric point 5.72. The result by GSDS (Gene structure display server) showed that the gene sequence of BmPTERP contains six exons and five introns. BmPTERP belongs to the superfamily of metallo-dependent hydrolases and contains specific hits of PTE. RT-PCR analysis revealed that BmPTERP was expressed in all the tissues tested, including gonad, midgut, hemocyte, fat body and silk gland. Real-time quantitative polymerase chain reaction analysis indicated that the relative transcript of BmPTERP in the normal midgut was approximately as 9.8 fold of that in the BmCPV-infected midgut.2 Molecular cloning and characterization analysis of 3-hydroxyacyl-coa dehydrogenase protein gene of silkworm, Bombyx mori (Bm3HCDH).Bm3HCDH consists of 1168 bp and contains a 83 bp 5’UTR and a 155 bp 3’UTR. A 930 bp open reading frame (ORF) encodes a 310 amino acid protein. The molecular weight of the putative HCDHBm was 33.6-KDa and isoelectric point 9.0. The result by GSDS (Gene structure display server) showed that the gene sequence of HCDHBm contains five exons and four introns. RT-PCR analysis revealed that Bm3HCDH was expressed in all the tissues tested, including gonad, midgut, hemocyte, fat body and silk gland. Real-time quantitative polymerase chain reaction analysis indicated that the relative transcript of Bm3HCDH in the normal midgut was 8.2 fold of that in the infected midgut.According to our research results of BmPTERP and Bm3HCDH, it is presumed that BmPTERP and Bm3HCDH might be involved in infection of silkworm with Bombyx mori cytoplasmic polyhedrosis virus. It provided not only new clues for investigating the molecular mechanism of BmCPV infection but also the theoretical basis for further study on the function of BmPTERP and Bm3HCDH of silkworm.

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CLC: > Biological Sciences > Molecular Biology > Genetic engineering (genetic engineering)
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