Qualitative and quantitative analysis of genetically modified pepper.

Area of Science:

• Agricultural Biotechnology
• Molecular Biology
• Food Science

Background:

• Genetically modified (GM) crops require reliable detection methods for regulatory and safety purposes.
• Developing specific and sensitive qualitative and quantitative PCR (Polymerase Chain Reaction) assays is essential for GM food analysis.
• Korea has developed GM pepper, necessitating validated analytical techniques for its identification and quantification.

Purpose of the Study:

• To develop and validate qualitative and quantitative PCR methods for detecting genetically modified (GM) pepper developed in Korea.
• To establish a reliable endogenous reference gene for accurate quantification of GM pepper.
• To assess the performance of the developed PCR method across various concentrations of GM pepper.

Main Methods:

• Utilized the capsanthin-capsorubin synthase (CCS) gene as an endogenous reference for PCR assays.
• Designed and tested primer pairs specific to the pepper endogenous CCS gene (102 bp amplicon) and the GM event (bar gene/Ti7 junction, 182 bp amplicon).
• Performed quantitative PCR using a TaqMan probe and a standard plasmid containing both endogenous and event-specific sequences for accurate quantification.

Main Results:

• The CCS gene primer pair (ccs-F/R) successfully amplified a 102 bp product from pepper DNA, with no amplification in 16 non-GM plant samples.
• Construct-specific primers amplified an 182 bp product from the GM pepper, confirming the presence of the integrated transgene.
• The quantitative PCR assay demonstrated accurate quantification of GM pepper in test samples ranging from 0.1% to 10% concentration.

Conclusions:

• Developed and validated robust qualitative and quantitative PCR methods for the detection and quantification of GM pepper.
• The CCS gene serves as a reliable endogenous reference for accurate GM pepper analysis.
• The established TaqMan-based quantitative PCR method is suitable for routine monitoring and regulatory compliance of GM pepper products.