I am doing knocking out for specific sequence by pX458 plasmid and I already did qPCR after 72h of transfection, I have used pX458 plasmid without gRNA as negative control but it seems that has the same effect like pX458 with gRNA because the relative amount of mRNA determined by PCR is similar, also when I extracted the RNA, RNA concentration was around 40 ng per micro letter for the treated cells, can any one help me please?
I performed qPCR on my samples in a setting similar to yours. But my K.O. samples were significantly different from negative controls.
Is there any statistically significant difference between your treated and untreated cells?
@Mohamed pirouzfar thanks for your answer, I have statical difference between the treated and non treated (control, cells only), but with the empty plasmid has effect more than the designer plasmid.
Can you tell me please which negative control you used?
Also the amplification cycle started at 32 for the treated and at 27 for the negative control, is that normal?
Heba
- Badges
- Science method