Focus on Microbiology Research - (Series 1)

Rapid, sensitive and specific detection of viable microorganisms is a fundamental requirement in many microbiological and quality control areas. DNA-based amplification techniques such as real-time PCR can provide the speed and specificity required. Living bacteria PCR is a next-generation technology for microbiology research that integrates cell survival information with the speed and specificity of PCR-based assays.

Living bacteria PCR is an innovative technology that distinguishes between living and dead microorganisms. The DNA insertion reagent, azide aziridine (PMA), was used in QIAGEN's live PCR system. PMA is a chemical molecule that distinguishes the survival of microorganisms at the DNA level. When a specific wavelength of laser is used to activate PMA, PMA will embed DNA and covalently bind to it. Amplification of the modified DNA in subsequent PCR reactions will be inhibited, resulting in a decrease in the amplification signal.

PMA does not pass through the complete biofilm structure. This means that DNA from viable microorganisms can be detected by PCR reactions because they are encapsulated by intact membrane structures and are not modified by PMA. The membrane structure of microbial dead cells loses its original protective function, and PMA can enter cells and modify its DNA. The DNA modified in the dead microbial cells is inhibited during the PCR reaction, resulting in a decrease in the amplification signal.

PMA has a strong positive charge, which means it cannot penetrate intact surviving cell membranes. It also has an anchoring group that is capable of undergoing an irreversible binding reaction with DNA under optical excitation at a specific wavelength. This feature allows PMA to compete beyond the binding of other DNA-embedded dyes to nucleic acids and is a fundamental component of QIAGEN's BLU-V® Viability product line.

The above information sources and QIAGEN official website!