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

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 (chk...

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Main Author:	Bnisallama, Faisal Fayez Saffah
Format:	Thesis
Language:	English
Published:	2022
Subjects:	QK900-989 Plant ecology
Online Access:	http://eprints.usm.my/58370/1/Bnisallama-24%20pages.pdf
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id	my-usm-ep.58370
record_format	uketd_dc
spelling	my-usm-ep.583702023-05-15T02:39:56Z Site directed mutagenesis of miRNA binding site on the 3′-UTR of choline kinase alpha gene 2022-09 Bnisallama, Faisal Fayez Saffah QK900-989 Plant ecology 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. 2022-09 Thesis http://eprints.usm.my/58370/ http://eprints.usm.my/58370/1/Bnisallama-24%20pages.pdf application/pdf en public masters Universiti Sains Malaysia Pusat Pengajian Sains Kesihatan
institution	Universiti Sains Malaysia
collection	USM Institutional Repository
language	English
topic	QK900-989 Plant ecology
spellingShingle	QK900-989 Plant ecology Bnisallama, Faisal Fayez Saffah Site directed mutagenesis of miRNA binding site on the 3′-UTR of choline kinase alpha gene
description	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.
format	Thesis
qualification_level	Master's degree
author	Bnisallama, Faisal Fayez Saffah
author_facet	Bnisallama, Faisal Fayez Saffah
author_sort	Bnisallama, Faisal Fayez Saffah
title	Site directed mutagenesis of miRNA binding site on the 3′-UTR of choline kinase alpha gene
title_short	Site directed mutagenesis of miRNA binding site on the 3′-UTR of choline kinase alpha gene
title_full	Site directed mutagenesis of miRNA binding site on the 3′-UTR of choline kinase alpha gene
title_fullStr	Site directed mutagenesis of miRNA binding site on the 3′-UTR of choline kinase alpha gene
title_full_unstemmed	Site directed mutagenesis of miRNA binding site on the 3′-UTR of choline kinase alpha gene
title_sort	site directed mutagenesis of mirna binding site on the 3′-utr of choline kinase alpha gene
granting_institution	Universiti Sains Malaysia
granting_department	Pusat Pengajian Sains Kesihatan
publishDate	2022
url	http://eprints.usm.my/58370/1/Bnisallama-24%20pages.pdf
_version_	1776101218191409152

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

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