Date of Award

11-2016

Document Type

Thesis

Degree Name

Master of Science (MS)

Department

Biology

First Advisor

Dr. Khaled Amiri

Second Advisor

Dr. Rabah Iratni

Third Advisor

Dr. Ghada Al-Kafaji

Abstract

Hammerhead RNA [HHR] sequence originally found in viroids is also detected in many species. HHR are often found in introns of different genes ranging from prokaryotes to

eukaryotes. HHR can form specific tertiary structure that make specific region susceptible to cleavage. However, the role of the HHR is not clear in other life forms. This study aims to

Investigate the role of HHR in the intron 6 of (RECK) gene. Our study shows that when blocking the HHR sequence in the intron6, the production of RECK protein declines which may be due to the effect of HHR in splicing and RNA stability which was seen by a decrease in the RNA levels of exons near to intron 6. Previous experiments designed to identify the significance of HHR sequence were addressing the self-cleavage of hammerhead RNA in vitro and comprised of partially constituted DNA construct expressed in cell culture. However, hammerhead RNA is a robust molecule and self-cleavage can take place even in biologically irrelevant context and therefore the current studies do not address its biological significance directly.

The aim of this study is to address the biological role of this sequence ex vivo. The study aims at blocking the endogenous mRNA with intronic hammerhead sequence in ex vivo. This study in principle should elucidate the biogenesis of mRNA whose HHR sequence is blocked. The biogenesis of mRNA will be assessed with RT-PCR. We will utilize bridge antisense (BNAs) RNA to block the HHR sequence in the mRNA. Furthermore, this study will investigate DNA pattern around the loci harboring HHR sequence. In this instance, efficient code will be written to search through the human genome data to identify locations that harbor patterns of the HHR sequence. Next, several thousands of bases upstream and downstream of this sequence will be extracted and analyzed to locate interesting patterns that may include gene, promoter regions and intronic regions.

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