There are additional polymorphisms surrounding my SNP of interest. Can I still use KASP chemistry, and how do I identify these surrounding polymorphisms in the SNP submission file?
KASP assays can include up to two other polymorphisms in the surrounding sequence (within 30 base pairs of the sequence of interest). These additional SNPs should be marked with their IUPAC codes, whilst the SNP of interest should be marked with square brackets surrounding it e.g. [A/G] or [R]. The assay design function in our Kraken software will design KASP primers that either avoid the surrounding polymorphisms or that incorporate degenerate / wobble bases into the primers to ensure that the primers are still able to bind efficiently to the DNA template.