This cell line you are analyzing, where you introduced a mutation via CRISPR/cas; is it homozygous or heterozygous?

If it were heterozygous one possible explanation is that the destabilization was so efficient that what you see in WB is what is being produced from the wild-type allele, therefore, what you are measuring post-cycloheximide is the half life of the product of the wild-type allele, and also, that whatever quantities are produced of the wild-type protein protein are sufficient to support miRNA biogenesis.

Hope it helps!

The mutation is homozygous (Sub cloning confirmed it).

Can you elaborate a bit on how you confirmed homozygocity by subcloning?

Also, how confident are you that the mutation does change the half life of the protein?

I used TA cloning kit and extracted 10 colonies. 9 gave me the exact same allele and the last one was not identified..regarding the protein change, well the mutated protein has decreased by 90% in WB analysis and i've ran 5 cycloheximide (100ug/ml) experiments in 6 timepoints (0,6,12,24,30,48)..normalization curves give the same trendline- flat.

Assuming your data for homozygosity is correct, one possible explanation is that your mutant has a defect somewhere on the protein biogenesis pathway, such as it does not fold correctly, or is not transported correctly etc (or associate with some needed component for folding or stability). So 90% of the synthesized protein is rapidly degraded whereas 10% did fold correctly and that subpopulation is stable like wild type.