Hi everyone
With the advent of CRISPR technology knocking out of genes has become a fairly easy and a week job. However I wonder to cope up with the permanent loss of vital genes the cells might adapt various counter balance mechanisms and only those clones that can overcome this loss finally survive during the selection process. In contrast the siRNA mediated Knockdown technology has its own drawbacks (off target effects, low efficiency, variability between experiments) but has some obvious advantages including absence of these counterbalance mechanisms as it is a sort of acute process when compared to knockout. Other thing with knockout is advantages and disadvantages of polyclonal vs single clone lines. Please share your experiences and thoughts on this issue.
Thanks
Dear Prafull,
You already have explained very well many of the pluses and minuses to each! We deal with this dilemma in my lab all the time. Not only is there RNAi and CRISPR/Cas9 to consider, but also chemical inhibitors (which have their own whole textbook of considerations).
Personally, I think its' essential to have multiple approaches. You need to look at the same question from different angles. For example, if you generate a knockout of protein X and see a phenotype, then you see the same phenotype with an inhibitor or RNAi, then you will be more confident that what you are seeing is not due to the drawback of either technique by itself. Like anything in science, a good conclusion is not drawn from a single experiment, but rather a confluence of evidence from different approaches that point to the same answer (and that then lead to novel testable predictions).
Dear Prafull,
Hope this helps :-)
http://www.genecopoeia.com/wp-content/uploads/2014/02/Technotes_Knockdown_vs_knockout.pdf
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