Electrochemical biosensor for genetic diagnosis based on catalytic activity of enzyme

Abstract

The detection of mismatched base pairs in DNA plays a crucial role in the diagnosis of genetic-related diseases and conditions, especially for early stage treatment. Among the various biosensors that have been used for DNA detection, EC sensors show great promise because they are capable of precise DNA recognition and efficient signal transduction. Par-ticularity specific application of enzymatic activity or electrochemical reagent for detection strategies have enabled for the development of highly sensitive, highly specific sensors making them attractive for the detection of small sequence variations. In the first strategy described in chapter 2, we developed a novel electrochemical method for SNP genotyping on the peptide nucleic acid (PNA)-modified electrode utilizing single-stranded DNA specific endonuclease. By using the strategy, we are able to success-fully detect various mutations including single base insertion, deletion, and substitution in BRCA1 gene for several real human samples from breast cancer patients. This study serves as a totally novel strategy for the electrochemical diagnosis of human genetic mutations ex-hibiting reliable discriminating power between wild type and mutant samples. The signifi-cance of the work derives from the following features. 1. This is the first report to utilize and combine the unique neutrality of PNA molecules and the delicate single-stranded DNA specific cleavage activity of nuclease S1 for the de-velopment of a new label-free electrochemical method for SNP genotyping. By ingeniously designing a novel strategy utilizing the electric charge property and distinctive cleavage ac-tivity, innovative label-free signaling detection was achieved and cumbersome extra-procedures for signaling were successfully eliminated, which are usually required for the previous electrochemical methods for SNP genotyping. 2. Since electrochemical method has many advantageous characteristics including sim-plicity, portability, and cos...