If I would like to knock down one particular gene in HeLa cells, would it be better to use CRISPR or RNAi. I don't want to create a stable cell line, just knock down one gene. What are the advantages or disadvantages of each system?
Hi Charles
That's a interesting question with long answers. Taking the long story short, if you do not want to make stable cell lines I will advice to use RNAi.
The CRISPr-based alternative to RNAi is to nased in the use a inactive mutant of Cas9 enzyme which will be transported to your DNA spot, blocking the RNA polymerase and decreasing the transcription rates (called iCRISPr). I never used this system, but from other colleagues I got the feeling that it is not as robust as RNAi. Other difference between the methods is that iCRISPr is a nuclear method, while RNAi will act at the cytoplasm.
It would be nice to have opinions of colleagues that already used both sytems for the knock-down of the same transcript.
Hope it helps. Best
Paco
Gene editing systems are more complex but the big advantage is that if you introduce something like an early premature stop codon, you can produce a true 'knockout' line (zero expression) rather than a 'knockdown' (still some expression though much lower than wild type).
Go to the kit Discussion group (https://groups.google.com/forum/#!forum/crispr).
Alot of this has been covered by the pioneers.
The most recent issue of Nature has a cool article on this subject by one o f the pioneers, Feng Zhang.
Soon CRISPR will supplant other RNAi based systems.
Se our recent paper on the world's first regulatory edit with CRISPR-Cas9 wherein we saw near complete knockdown of a target gene in a living animal
Article CRISPR-Cas9 Genome Editing of a Single Regulatory Element Ne...
You can go through this article. possibly will help you.
http://www.genecopoeia.com/wp-content/uploads/2014/02/Technotes_Knockdown_vs_knockout.pdf
Hello.
All friends have pointed to the good point and useful article, That all are correct.
But besides this, note that the system CRISPR requires high skill and expertise, especially for design gRNA. If you have this ability would be useful to use this method
additional blog articles that evaluate CRISPR and RNAi when it comes to off-targeting:
http://blog.sitoolsbiotech.com/2017/04/crispr-what-can-go-wrong-and-how-to-deal-with-it/
http://blog.sitoolsbiotech.com/2017/07/sirna-vs-shrna-applications-and-off-targeting/
Essentially, its important to use an array of techniques to thoroughly establish gene function!
RNAi is going to be cheaper and have higher efficiency. We've made a kit for HeLa (https://altogen.com/product/hela-transfection-reagent-cervix-adenocarcinoma-cells/) with siRNA-based gene silencing validated to 90% efficiency. The results will be transient, in contrast to the stable changes caused by CRISPR/Cas9, however for most research such transient silencing is sufficient.
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