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Hello tramelliwe I can't really speak on whether the two peaks are common, but I can say that after doing a lot of amplicon sequencing I've had a lot of off-target amplification. Depending on what you're sequencing it can be expected and it happens even more with degenerate primers. Have you looked to see if any of your primers match up with that region? I can also check in with some friends that have more experience with this type of work. -
Thank you for your answer Ben3. Maybe I should clarify that my amplicon covers both peaks. I did check the specificity, but even with multiple mismatches, my primers do not bind other places on that locus. I did not do the library prep myself, maybe at that stage the fragments covering that zone were lost somehow?
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CRISPR/Cas9 sparked the gene editing revolution for both research and therapeutics.1 But this system still showed severe issues that limited its applications. The most prominent were the heavy reliance on PAM sequences, delivery limitations, double-stranded breaks that prompt unintended edits and cell death, and editing inefficiency (both in targeting and in knock-in reliability).
Despite this, “CRISPR helped turn genome editing from a specialized technique into...07-31-2026, 11:01 AM
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