Electrochemical Primer Extension for the Detection of Single Nucleotide Polymorphism Using Dideoxynucleotides Labeled with Polyoxometalates

Entity: Universitat Rovira i Virgili (URV)

Confidenciality: No

Education area(s): Nanociència, Materials i Processos - Tecnologia Química de Frontera 

Title in different languages: Electrochemical Primer Extension for the Detection of Single Nucleotide Polymorphism Using Dideoxynucleotides Labeled with Polyoxometalates

Abstract: Single Nucleotide Polymorphism (SNPs) are important genetic markers that can be helpful for identifying predisposition to a disease, patient stratification in the future paradigm of personalized medicine as well as an advanced forensic tool. Following identification of the location of SNPs using next generation sequencing technologies, primer extension reactions can be used for rapid identification of which SNP is present at the identified SNP site using dideoxynucleotides (ddNTPs) that will elongate a DNA double strand by just one base exactly at the SNP site. A novel approach of the PEX reaction previously introduced by our group is Electrochemical Primer Extension (EPEX), where dideoxynucleotides (ddNTPs) labelled with different electrochemical markers were tested. In this work, for the first time ddNTPs labelled with Keggin and Dawson polyoxometalates (POMs) are effectively incorporated using the EPEX reaction to elongate immobilized primers using the cardiomyopathy MYH7 gene as a model system. We successfully detected four different SNPs, representing each of the bases, using differential pulse voltammetry to detect the incorporated redox-labelled ddNTPs. The polymerase chain reaction was carried out to amplify a base pair region containing the four different bases, as a means of stimulating real patient samples. Once the target DNA products from asymmetric PCR were obtained, the single stranded DNA were tested in the EPEX reaction obtaining 100% of correct base incorporation and SNP detection for the Dawson POM. This study is an important development to achieve a cost-effective platform for detecting SNPs at identified locations.

Subject: Enginyeria química

Academic year: 2015-2016

Language: Anglès

Work's public defense date: 2016-09-08

Subject areas: Chemical engineering

Student: Uribe Uribe, Laura

Work's codirector: Ortiz, Mayreli

Department: Enginyeria Química

Creation date in repository: 2017-01-13

TFM credits: 24

Keywords: Single Nucleotide Polymorphism, Dideoxynucleotides, Polyoxometalates.

Title in original language: Electrochemical Primer Extension for the Detection of Single Nucleotide Polymorphism Using Dideoxynucleotides Labeled with Polyoxometalates

Access Rights: info:eu-repo/semantics/openAccess

Project director: O'Sullivan, Ciara