| Summary: | 博士 === 高雄醫學大學 === 臨床醫學研究所 === 106 === (一) Long noncoding RNAs (lncRNAs) have various functions, including chromatin remodeling and the regulation of gene expression at both transcriptional and posttranscriptional levels. However, few lncRNAs have been comprehensively studied, with the majority being uncharacterized. In this study, we established a bioinformatics pipeline to identify novel lncRNA sequences similar to the 3′-untranslated regions (3′-UTRs) of protein-coding genes. These pairs of lncRNAs and coding genes contained the same miRNA target sites; the lncRNA CR933609 matched the 3′-UTR of INO80D mRNA. The expression levels of CR933609 and INO80D were significantly decreased in non-small-cell lung cancer (NSCLC) and other cancer tissues. The expression levels of both CR933609 and INO80D decreased in CR933609 knockdown NSCLC cells, but only INO80D expression levels decreased in INO80D knockdown cells. We proved that there are independent promoters in CR933609 and INO80D. We also found that expression levels of INO80D were downregulated by endogenous miRNA-5096 in A549 cells, but not in CR933609-overexpressing A549 cells. Furthermore, the lncRNA CR933609 acts as a decoy to protect INO80D from downregulation by miRNA-5096 in NSCLC cells. We established a protocol to identify novel lncRNAs in the 3′-UTR and proved the existence of novel lncRNAs.
(二) Cytochrome P450 (CYP) 2C8 is the principal enzyme responsible for the metabolism of arachidonic acid and various drugs and influences drug-drug interactions and some associated diseases. Large interindividual differences in CYP2C8 enzymatic activity and several nonsynonymous genetic variations have been reported in different races. Therefore, how to identify CYP2C8 polymorphisms efficiently for genotyping in different populations is very important. A high resolution melting (HRM) analysis was used to characterize the CYP2C8 polymorphism. Genomic DNA was extracted from peripheral blood samples from 95 normal individuals in Taiwan. Nine exons of the CYP2C8 gene were screened by HRM analysis. All results were confirmed by direct DNA sequencing. Five new single nucleotide polymorphisms (SNPs) were found in this study; two SNPs [1189G>A (D397N) and 1230C>T (G410G)] were in exon 8 and the others [1312G>C (E438Q), 1497T>C (A499A) and 1677delT (559delL)] were in exon 9. The 1497T>C (A499A) was the most common variant with an allele frequency of 20.53% but without amino acid substitution. HRM analysis is a fast, reliable, accurate and cost-effective screening method for gene mutations, even very similar cDNA sequences with 83% identities, compared with CYP2C8 and CYP2C9.
|
|---|
