Date Written

2017

Document Type

Honors Paper

Degree Name

Biochemistry and Molecular Biology-BS

Department

Biochemistry and Molecular Biology

Advisor

Dr. John Tansey

First Committee Member

Dr. Jennifer Bennett

Second Committee Member

Dr. Margaret Koehler

Keywords

Perilipin 5, Alternative Splicing, Northern Blot, Diabetes, Gene Expression, Lipid Droplet

Subject Categories

Biochemistry | Molecular Biology

Abstract

Alternative splicing occurs throughout the human genome, leading to multiple proteins from a single gene. The resulting proteins can be nearly identical or vastly different in how they function. Alternative splicing within the perilipin family has been observed in perilipins 1 and 3, giving rise to proteins with varying functions. In the course of our studies on perilipin 5, an immunoblot signal was observed that corresponded to an approximately 35 kDa protein. Western blot analysis of this protein has revealed its expression in C2C12 cultured myoblasts and in heart and liver mouse tissue. Further analysis of perilipin 5 cDNA through reverse transcriptase PCR provided evidence that this protein is a result of alternative splicing occurring in perilipin 5 that we have termed perilipin 5b. The objective of the current study is to develop DNA based probes for use in northern blot analysis of perilipin 5. The genomic DNA sequence of perilipin 5 was analyzed to identify areas where splicing events could occur, and probes were designed accordingly to detect the different variants of perilipin 5. In the analysis of different mouse tissue samples, these probes have been used to detect which of the perilipin 5 splice variants are being expressed under different feeding conditions. We hypothesize that probes made to detect the 5’ end of perilipin 5 will detect both variants, while probes for the 3’ end will selectively bind to one variant.

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