Question: Why are the enrichment primers designed close to the 5' end of the C-region in the V(D)J assay?
Answer: The reason that the enrichment primers are located near the 5' end of the C-region in the V(D)J assay is to capture only the full length V(D)J sequence information, while keeping the enriched PCR fragments as short as possible. If the primers were located closer to the 3' end of the transcript, this would result in more constant region sequencing coverage, which is not ideal as this is not the information we are interested in capturing. In addition, diversity is needed in the 5' region upstream of the primer in order to distinguish various isotypes.
The enrichment primers are designed to balance these two aspects: sufficient diversity and proximity to 5' end.
Products: Single Cell Immune Profiling