Presentation Title

Cloning and Characterization of p53 DNA Binding Domain

Start Date

November 2016

End Date

November 2016

Location

HUB 302-155

Type of Presentation

Poster

Abstract

P53 is a tumor suppressor protein that binds to the DNA and is responsible for regulating cell growth. Mutations of p53 are associated with numerous types of cancers in patients worldwide. Mutants of p53 affect not only its structure, but its original function, causing p53 to gain oncogenic functions. The focus of this research is to clone the DNA binding domain (DBD) from the wild type p53 as well as its most common mutants to characterize the p53 DBD. It is proposed that when missense mutations are present within the p53 DBD, p53 will destabilize, leading to its aggregation within the cell. We used molecular cloning techniques such as PCR to isolate the DBD from p53. We also utilized mammalian cell viability assays such as MTT and cytotoxicity to determine the effects of p53 on human cells. PCR results showed that DBD was amplified from full length p53 and cloned into expression vectors pET28 and pcDNA3.1. In our preliminary results both the MTT and cytotoxicity assays showed that WT p53 decreases the viability of mammalian cells. Therefore, we successfully cloned the DBD from p53 and are in the process of characterizing the effects of mutant p53 on cell viability and aggregation propensity.

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Nov 12th, 1:00 PM Nov 12th, 2:00 PM

Cloning and Characterization of p53 DNA Binding Domain

HUB 302-155

P53 is a tumor suppressor protein that binds to the DNA and is responsible for regulating cell growth. Mutations of p53 are associated with numerous types of cancers in patients worldwide. Mutants of p53 affect not only its structure, but its original function, causing p53 to gain oncogenic functions. The focus of this research is to clone the DNA binding domain (DBD) from the wild type p53 as well as its most common mutants to characterize the p53 DBD. It is proposed that when missense mutations are present within the p53 DBD, p53 will destabilize, leading to its aggregation within the cell. We used molecular cloning techniques such as PCR to isolate the DBD from p53. We also utilized mammalian cell viability assays such as MTT and cytotoxicity to determine the effects of p53 on human cells. PCR results showed that DBD was amplified from full length p53 and cloned into expression vectors pET28 and pcDNA3.1. In our preliminary results both the MTT and cytotoxicity assays showed that WT p53 decreases the viability of mammalian cells. Therefore, we successfully cloned the DBD from p53 and are in the process of characterizing the effects of mutant p53 on cell viability and aggregation propensity.