A laboratory switches its real-time PCR assay for a DNA virus from a hydrolysis probe chemistry to an intercalating dye such as SYBR Green. Compared with the probe-based assay, the main technical drawback of the dye-based assay is:
- A It fluoresces regardless of which double-stranded DNA is present, so non-specific products and primer dimers also give signal ✓
- B It cannot amplify targets longer than 150 base pairs
- C It cannot be used with a melting curve analysis
- D It requires a separate labelled probe for every run
Correct answer: A. It fluoresces regardless of which double-stranded DNA is present, so non-specific products and primer dimers also give signal
Explanation
SYBR Green binds any double-stranded DNA generated in the reaction, so primer dimers and non-specific amplicons fluoresce exactly like the intended product, reducing specificity. This is why dye assays require post-run melting curve analysis to confirm product identity. Option C is wrong because melt curves are a feature of dye chemistry, and D describes probe systems, not dye systems.
Reference: Ananthanarayan and Paniker's Textbook of Microbiology, 11th ed.
High-yield for: NEET PGINI-CETNExTFMGEUSMLEPLABMRCP
Written and medically reviewed by the StethoPrep medical team.