What is FAM in qPCR?

What is FAM in qPCR?

FAM, or 6-carboxyfluorescein, is a fluorescent dye commonly used in quantitative Polymerase Chain Reaction (qPCR) assays. It is a reporter molecule that allows for the detection and quantification of a specific DNA sequence in real-time PCR applications. FAM is attached to a probe that hybridizes to a complementary DNA sequence within the target DNA. When the probe is intact, the fluorescence of FAM is quenched. However, during the PCR process, the probe is degraded by the 5' to 3' exonuclease activity of Taq polymerase, releasing the FAM dye and allowing it to fluoresce. This fluorescence can be measured in real-time, providing quantitative information about the initial amount of the target DNA.

Key Features of FAM in qPCR:

• Fluorescent Reporter: FAM serves as a fluorescent reporter dye, emitting a green fluorescence when excited, typically at a wavelength of 495 nm and emitting at 520 nm.
• Quantification: The intensity of the fluorescence signal is proportional to the amount of target DNA, allowing for quantitative analysis.
• Sensitivity: FAM-labeled probes provide high sensitivity and specificity in detecting target DNA sequences.
• Versatility: FAM is compatible with most qPCR instruments and can be used in multiplex assays with other dyes, provided they have different emission spectra.

Applications of FAM in qPCR:

• Gene Expression Analysis: Quantifying mRNA levels of specific genes to study gene expression under various conditions.
• Genotyping: Identifying genetic variations, such as single nucleotide polymorphisms (SNPs).
• Pathogen Detection: Detecting and quantifying viral, bacterial, or other microbial DNA/RNA in samples.
• Copy Number Variation: Assessing variations in the number of copies of a particular gene across different genomes.