shRNA depletion and enrichment screening
Posted 13 March 2011 - 04:56 PM
I've also been reading about positive selection versus negative selection shRNA screening. Are these names the same as saying the shRNA has gone through enrichment or depletion screening ?
Can someone give me information on the web to read involving shRNA enrichment and depletion studies, I just can't find any it seems!
I assume the above questions can also be applicable to other RNAi applications like siRNAs and miRNAs as well?
Posted 13 March 2011 - 06:10 PM
shRNA library screening works this way: a library of shRNAs that targets all genes or a group of genes are used to infect cells at a certain MOI so that one cell will only be infected by one viral particle. If the researcher is interested in identifying certain genes whose knockdown will cause a particular phenotype, eg, M phase arrest. After the shRNA infection, the researcher would enrich M phase cells by FACS analysis and recover the shRNA from the enriched cells. By identifying which shRNA(s) are enriched in the M phase cells, the target gene (one or more) that promote M phase is then identified. This make perfect sense if the knockdown causes phenotypes which could be used to enrich the infected cells. If the knockdown causes depletion of infected cells, recovering the responsible shRNA is very complicated involving using many control treatments.
This page gives pretty good definitions:
Posted 20 March 2011 - 04:10 PM