Molecular screening using non radioactive differential display technique in Malay Kelantanese patients with peptic diseases

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Date
2005
Authors
Wan Taib, Wan Rohan!
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Abstract
Peptic diseases are the most common chronic diseases of adulthood and proven to have a substantial multifactorial inherited components. Genetic influences play some role in the predisposition to both forms of ulcers (gastric and duodenal ulcer). A small proportion of chronic gas~c ulcers are susceptible to be transformed into malignancy. The possible somatic mutatio~s that ~e place have not been extensively studied. The discovery of some genetic changes at the vicinity of the chronic benign inflammatory lesions is important in relation to the elucidation of the carcinogenesis of gastric cancers. The general aims of this study were to screen for differentially expressed genes in peptic diathetic patients and to apply a technique of non radioactive differential display analysis (DDRT-PCR). DDRT-PCR has been shown to be highly effective in identifying sequences that are differentially expressed in various cell types and this technique makes it possible to obtain reproducible result and efficiently identify specific mRNAs. Twenty tissue sample biopsies of gastric mucosa of the antrum were collected from peptic diathetic patients at Endoscopy unit. Total RNAs were extracted by using RNA extraction kit (RNeasy Mini Kit, Qiagen). The DDRT-PCR analysis was performed by a 2- step method which were reverse transcription and polymerase chain reactions (RNAimage Kit 1, GenHunter). Six percent denaturing Polyacrylamide Gel Electrophoresis (PAGE) was carried out in order to obtain the size of separation of eDNA fragments and visualized by silver staining. Once differentially expressed mRNAs were identified, the corresponding cDNAs were eluted from the band of the gel and reamplified. The sequence of cDNAs were determined using an ABI Prism DNA XVI Sequencer. The sequences were searched for its homology using GenBank. databases provided by National Institutes of Health (NIH, USA). Two differentially expressed genes were identified, namely, ubiquinol-cytochrome c reductase complex (Complex III) gene and ribosomal protein L27a gene in gastritis tissue compared to normal gastric tissue. The expressed genes can be analyzed to determine their involvement in the pathogenesis of peptic diathesis. The determination of these genes will be used to study whether similar genetic derangement occur in gastric cancers in the future. This knowledge will enhance the understanding of carcinogenesis of chronic inflammatory lesions.
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Malay Kelantanese , Peptic diseases
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