Question

qPCR using TaqMan probes have the advantage of being very specific for the harget species, even when closely related species could also be amplified (e.g., African vs. Indian elephants). TaqMan probes are often 30bp in length. If you had a genome of 3 billion base pairs long: a) provide how many theoretical times that probe should bind to the genome b) explain how the TaqMan probe makes qPCR so much more specific than just having 2 primers to amplify a locus.

Answer 1

Please find the answers below:

Answer a) According to the information, the probe has a length of 30 bp and the target DNA has nearly 3 billion base pairs. Thus, provided that the probe can randomly bind to the genomic DNA, there can be 3*10⁷/30 ways or 10⁶ or 1 million ways.

Answer b) The TaqMan probes are highly specific in nature and are designed in accordance with the exact genetic make up of the species, even when two organisms are very closely related to each other. In this regard, it must be carefully noted that there is a very minute, but significant, difference among the genetic sequences for African vs Indian elephant and hence, this minute genetic difference is exploited to make a very specific probe against the gene and make them bind in the qPCR reaction.