FAQ: Why do I see products of the wrong size?

There could be several reasons:
  1. The presence of a PCR product larger than expected is often due to the contamination of genomic DNA. Treat with DNase I prior to cDNA synthesis (5).
  2. The presence of a PCR product of the correct size in the -RT negative control is either due to contaminating genomic DNA or carryover PCR product. Use separate areas for reaction assembly and product analysis. Use primers spanning an exon-exon boundary.
  3. Non-specific PCR products can be eliminated by optimizing PCR reactions. This involves the following: (1) check your primers with a computer program, (2) increase annealing temperatures in 1°C increments, and (3) lower primer concentration to 75 nm.