Site directed mutagenesis of miRNA binding site on the 3′-UTR of choline kinase alpha gene

Bnisallama, Faisal Fayez Saffah (2022) Site directed mutagenesis of miRNA binding site on the 3′-UTR of choline kinase alpha gene. Masters thesis, Universiti Sains Malaysia.

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MicroRNA largely controls gene expression by attaching to messenger RNA (mRNA) in the cell cytoplasm. Instead of being promptly translated into a protein, the targeted mRNA will either be destroyed and its components recycled, or it will be retained and translated later. Choline kinase alpha (chka) overexpression is a clinical sign of diseased tissues and malignant cells. MicroRNAs (miRNAs) are effective posttranscriptional regulators of gene. Studies by our team showed that these three miRNAs (miR-876-5p, miR-367-3p and miR-32-5p) downregulated the expression of chka gene. However, the binding sites of these miRNAs on the 3'-UTR of the chka gene have not been verified. This study aimed to mutate the binding sites of these miRNAs for subsequent verification by a luciferase assay. In this study, we performed PCR site-directed mutagenesis on the miR-367-3p (GAAGCAGAAAT ATAGTGCAATA) from nucleotides (nt) 1817-1825 binding sites in chka and miR-876-5p (GAG TGTAGCTGTG AAATCCA) binding site from nucleotides (nt) 2573-2581 binding sites and verified the mutation by DNA sequencing. In vitro work, after mutagenesis step, the PCR products were sent for sequencing, however the results were not satisfactory.

Item Type: Thesis (Masters)
Uncontrolled Keywords: miRNA, choline kinase alpha gene
Subjects: Q Science > QK Botany > QK900-989 Plant ecology
Divisions: Kampus Kesihatan (Health Campus) > Pusat Pengajian Sains Kesihatan (School of Health Sciences) > Thesis
Depositing User: Mr Abdul Hadi Mohammad
Date Deposited: 07 May 2023 04:03
Last Modified: 15 May 2023 02:39

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