RNA interference screening demystified

normally arrayed in 96-well or 384-well plates

can combine multiple siRNAs targeting different sequences in the same gene in one well (SMARTPools)

can be purchased in “ready-to-transfect” aliquots or in larger amounts that require replating

can also be arrayed on slides

Consistent quality of reagents

Ease of use and readily transfectable

Chemical modification of siRNA can limit off-target effects

Finite resource

Target cells need to be transfectable

Relatively short period of silencing

Not suited to pooling strategies

plasmid DNA arrayed in 96-well or 384-well plates for transfection

viral particles in multi-well plates (one shRNA per well)

pools of plasmids or viral particles

Renewable resource

Viral vectors enable silencing in difficult-to-transfect cells

Suited to pooled screens as well as arrayed screens

Stable integration of silencing machinery into host cell genome enables longer-term silencing

Varying vector formats available that allow inducible silencing, tracking of silencing machinery, etc

Reduced ease of use: require preparation of plasmid DNA and, in the case of viral-based libraries, viral packaging

Viral use often requires biological containment

Pooled screens require significant deconvolution