Real-time PCR/qPCR (CAT#: STEM-MB-0185-WXH)

The principle of real-time PCR relies on the use of fluorescent dye. In general, the principle of the present method is: “The amount of the nucleic acid present into the sample is quantified using the fluorescent dye or using the fluorescent-labeled oligos.” When a dye or probe binds with the target template, it releases a fluorochrome which resultantly emits fluorescence for the detector to detect. The detector captures a signal as a positive template amplification.
Two types of chemistry are available for the real-time quantitative PCR:
1.DNA binding dye (Intercalating dye-based method)
2.Sequence-specific probe (Hydrolysis Probe-based detection method)

• Quantification of DNA, RNA and gene expression.
• Disease diagnosis and management.
• Animal and plant breeding: Gene copy number.
• Used in food safety, food spoilage and fermentation.
• For the microbial risk assessment of water quality and in public health protection.
• Detection of genetically modified organisms.
• Detection of phytopathogens.
• Clinical quantification and genotyping: Viral quantification.

1.Denaturation
2.Annealing
3.Extension

Taqman Probe: It is a hydrolysis probe which bear a reporter dye, often fluorescein (FAM) at its 5’ end and a quencher tetramethylrhodamine (TAMRA), attached to the 3’ end of the oligonucleotide.
SYBR Green: This is a dye that emits prominent fluorescent signal when it binds at the minor groove of DNA, nonspecifically. Other fluorescent dyes like Ethidium Bromide or Acridine Orange can also be used but SYBR Green is better used for its higher signal intensity.