FAQ: The sgRNA is not running as a single band on the gel – why is this happening?
We recommend running the RNAs on a TBE-Urea (denaturing) gel under
denaturing conditions. The sgRNA is highly structured and if not fully
denatured may run as multiple bands on a gel and may not run at the
correct size as compared to a ssRNA ladder such as the Low Range ssRNA
Ladder (NEB# N0364). If smearing is present, this may indicate
contamination with an RNase. We recommend wearing gloves and using
nuclease-free tubes and filter tips