How much DNA template should I use in my KASP genotyping reactions?
The minimum final DNA concentration that LGC recommend in KASP genotyping reactions is 2.5 ng / µL. For example, if you were preparing a 10 µL reaction consisting of 5 µL of DNA and 5 µL of genotyping mix (KASP Master mix + KASP Assay mix) then the input DNA would need to be at 5 ng / µL to ensure a final concentration of 2.5 ng / µL.
This value is based on the human genome size (~3000 Mbp). If the genome size of your study organism is larger than human, you will need to adjust final DNA concentration (and hence input concentration) accordingly. (Please note that we do not recommend reducing the input DNA concentration for genomes smaller than human).
For genomes larger than human, a higher concentration of DNA is required. To calculate this, divide the genome size of your organism by the size of the human genome (3000 Mbp), and use the resulting number to multiply the final concentration of DNA that should be used in your KASP reactions.
e.g. Triticum aestivum (wheat): 15966 Mbp
15966 Mbp / 3000 Mbp = 5.3
You will need a final DNA concentration that is 5.3 times more concentrated = 2.5 ng / µL DNA x 5.3 = 13.25 ng / µL final concentration.