Paper Information

Title: 

BCR-ABL SILENCING BY USING SPECIFIC SMALL INTERFERENCE RNA (SIRNA)

Type: POSTER
Author(s): JAVADI H.R.*,ZAREI A.
 
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Name of Seminar: IRANIAN CONGRESS OF PHYSIOLOGY AND PHARMACOLOGY
Type of Seminar:  CONGRESS
Sponsor:  PHYSIOLOGY AND PHARMACOLOGY SOCIETY, MASHHAD UNIVERSITY OF MEDICAL SCIENCE
Date:  2007Volume 18
 
 
Abstract: 

Introduction: Chronic myelogenous leukemia (CML) is a human malignancy marked by the presence of a distinct cytogenetic abnormality which results from a translocation between chromosomes 9 and 22, t(9;22), known as Philadelphia chromosome. This translocation causes aberrant expression of Bcr-abl, a constitutively active tyrosine kinase that has been linked to the pathogenesis of CML. Bcr-abl is thought to promote malignant transformation by altering cellular adhesion properties, stimulating mitogenic signaling pathways, and inhibiting programmed cell death (apoptosis). Bcr-Abl prevents apoptosis through inhibition of mitochondrial cytochrome C release or inhibition of caspase activation after the release of cytochrome C. Therefore it appears that Bcr-Abl silencing can induce apoptosis in CML cells. The aim of this study was to silence Bcr-Abl gen by using a specific siRNA (19-22 mer nucleotide) in cultured K562 cell line.
Methods: by using mRNA sequence of Bcr-abl from NCBI, various siRNA targeting Bcr-abl mRNA were designed and blasted in NCBI, then the sutiable shRNAs was choosed and synthesised. then the suitable shRNA inserted to pRNAH1.1/Neo vector by using BamHI and HindIII restriction enzymes. Recombinant vector delivered to DH5α (E.coli) and developed. The correct transmission of expression vector was confirmed by using of restriction enzymes and electrophoresis. Expression vectors which were isolated from DH5α (Ecoli), transfected to K562 cell lines by using of electroporation procedure and incubated in culture media. Gene silencing was investigated by monoclonal Ab against Bcr-abl gene product.
Results: restriction digest confirmed correct inserted Gene product. The extent of gene silencing by western blot investigation was shown significant reduce in gene products.
Conclusion: This results, suggest that siRNA is a suitable for gene silencing hn cancer and viral.

 
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