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Studies on the apoptotic effect of canine distemper virus (CDV) genes in tumor cell-lines

KrishiKosh

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Title Studies on the apoptotic effect of canine distemper virus (CDV) genes in tumor cell-lines
 
Contributor Deka, Dipak
 
Subject Canine distemper virus
virus isolation
wild-type
molecular characterization
apoptosis
 
Description Canine distemper (CD), caused by canine distemper virus (CDV) is a highly contagious disease that infects a variety of carnivores. Although only one CDV serotype has been reported worldwide, lot of genetic variation in the virus genome particularly in the haemagglutinin (H) gene has been observed in CDV strains of different regions which are thought to be one of the causes of vaccine failure. In this study, a total of 50 ocular and nasal swab samples from CDV suspected dogs when processed for viral RNA detection, desired amplicons of 268bp could be detected in three cases. The CDV positive samples were then subjected to virus isolation first in primary dog lymphocyte culture and then in MDCK cells which were again harvested, passaged and viral RNA was confirmed in cell culture harvest by diagnostic RT-PCR. The individual CDV genes of a local isolate viz. full length fusion protein (F), phosphoprotein (P) matrix protein (M) and partial coding sequence of haemagglutinin (H) and largeprotein (L) were amplified, cloned and then sequenced. Both nucleotide and amino acid sequence alignment of all the genes revealed a higher sequence homology of the Indian isolate with that of the strains from Switzerland, Hungary and Germany; and a lower with that of the vaccine strains like Ondersteport, CDV3 and Convac. More sequence variation was observed in H and F genes as compared to the P and M genes. Predicted potential N-linked glycosylation sites in H, F, M and P proteins of the local isolate were similar to the previously known wild-type CDVs but different from the vaccine strains. The Indian CDV formed a distinct clade in the phylogenetic tree clearly separated from the previously known wild-type and vaccine strains. Subsequently, the apoptotic potential of CDV as well as its individual proteins was evaluated by infecting HeLa cells (cancerous cells) with the local isolate and also by transfecting the cells with the individual CDV genes (F, M and P) cloned in an expression vector. All the assays i.e. DNA laddering, MTT assay, TUNEL assay and Apo-Tox Glo kit for caspase 3/7 activity, revealed that CDV induced apoptosis in cervical tumor derived cell-line (HeLa cells). Among the CDV genes/proteins, M protein could induce apoptosis to some extent in the transfected HeLa cells as compared to other proteins (P and F) clearly indicating its apoptotic potential.
 
Date 2016-08-25T11:37:41Z
2016-08-25T11:37:41Z
2015-11-13
 
Type Thesis
 
Identifier http://krishikosh.egranth.ac.in/handle/1/73732
 
Language en
 
Format application/pdf