What is the reason for split peaks appearing in between dispensations on my pyrosequencing pyrogram?

The PyroMark cartridge needle can be blocked or damaged. Clean the cartridge or exchange with a new one. Check for correct reagent cartridge and cartridge method used in the run. Check if the reagent cartridge cover was closed properly. Make sure that the cartridge was dry after cleaning because nucleotide droplets might be caught at the needle tip and might fall down at any time, or exchanged.