Semi-quantitative detection of gene expression using bisbenzimide dye

Area of Science:

• Biotechnology
• Molecular Biology
• Electrochemistry

Background:

• Gene expression analysis is crucial for understanding cellular function and disease.
• Conventional methods like agarose gel electrophoresis have limitations in sensitivity and throughput.
• Developing rapid, sensitive, and accurate gene detection methods is essential.

Purpose of the Study:

• To develop and validate an electrochemical biosensor for detecting gene expression.
• To compare the sensitivity and accuracy of the electrochemical biosensor with conventional agarose gel electrophoresis.
• To assess the RAGE gene expression levels in HeLa and HepG2 cell lines.

Main Methods:

• An electrochemical biosensor utilizing a disposable printed chip and Hoechst 33258 dye was employed.
• Linear sweep voltammetry was used to quantify gene expression based on anodic peak current.
• RNA was reverse transcribed to cDNA, amplified by PCR, and then detected by either electrophoresis or the electrochemical biosensor.

Main Results:

• The electrochemical biosensor demonstrated acceptable precision for both β-actin and RAGE gene detection.
• The biosensor method showed high correlation and no significant difference compared to agarose gel electrophoresis.
• The electrochemical biosensor was more sensitive, detecting as low as 10 copies of the RAGE gene, compared to 10^4 copies for electrophoresis.
• HeLa cells exhibited approximately twofold higher RAGE gene expression than HepG2 cells.

Conclusions:

• The developed electrochemical biosensor is a sensitive and reliable tool for gene expression analysis.
• This method offers advantages over conventional electrophoresis in terms of sensitivity and potential for high-throughput screening.
• The biosensor, modified with Hoechst 33258, shows promise for detecting gene expression in various cell types and research applications.