Dear all,
I have been trying to knockdown using siRNA (duplex) targeted against my protein of interest (# 2 lane shown in the image) and a random non-target siRNA ((# 1 lane shown in the image) treated cell as negative control and non-treated normal cell (# 3 lane shown in the image). I don't see silencing of the gene as could be seen in lane 2. Respective actin loading control is shown below the bands of interest. Currently, I am using 20 nM siRNA with interferring reagent protocol provided from thermofischer. If anyone can suggest any protocol that I can try, will be greatly helpful.
Thank you
best
Prem
Hi Prem,
It seems your knockdown experiments does not work well. There are some reasons:
1- Your siRNAs are not target the right target gene.
2- Your siRNAs could not enter inside the target cells.
3- Your siRNAs were degraded during infection.
If possible, you can try to transform your cells by a siRNA-expression plasmid. This plasmid can express the siRNA which bind your target mRNA.
Good luck
Trung
Not all proteins are regulated by by transcript abundance. If the protein is rarely degraded, then it can time a long time for levels to drop.
- Badges
- Science topic
- Similar topics
- Phytochemistry
- Extracts