What do first round primers in Nested/Semi-Nested PCR aim to target?

In a Nested or Semi-Nested PCR, the first round primers are designed to target longer products within genomic cDNA. This process is based on the need to amplify a specific DNA region that may be present in low quantities among a complex mixture of nucleic acids. The first round of amplification serves to generate a larger initial product, which can enhance the sensitivity of the reaction.

Specifically, these longer products provide a broader template for the subsequent round of PCR with nested primers, which are designed to be more specific and amplify a subregion of the initial product. This two-step approach improves specificity and reduces the chances of non-specific amplification.

The other options are less relevant in this context. While shorter fragments could be targeted by different primer sets, the focus of the first round in a nested approach is to capture a larger region of interest. Fragments of RNA are not the direct target of PCR, as PCR is specifically used for amplifying DNA. Lastly, targeting unrelated sequences would not serve the purpose of generating amplification for a specific target, which is essential for the success of nested PCR methodologies. Therefore, focusing on long products within genomic cDNA aligns with the objective of enhancing specificity and sensitivity in the amplification process.