| Strengths |
-
•
Can use diverse cell lines
-
•
High transfection efficiency of adherent cells
-
•
Increased sensitivity: arrayed format permits selection of a gradation of phenotypes
-
•
Library key permits rapid gene identification
-
•
Arrayed format permits screening for viral budding/production
-
•
Can perform image-based screens and investigate cell biology phenotypes
-
•
Creates hypomorphs permitting many essential genes to be screened
-
•
Readily validated using reagent redundancy
-
•
Short-term screens < 10 days
|
-
•
Can use diverse cell lines
-
•
Viral transduction works better for suspension cells
-
•
Good format for suspension cells
-
•
Long-term screens (> 10 days)
-
•
Lower cost than siRNA once the shRNA library is purchased
|
-
•
Finds receptors, entry factors, and associated genes
-
•
High specificity: less false positives
-
•
Generates null phenotype
-
•
Long-term screens (> 10 days)
-
•
Low cost to perform survival screens
|
-
•
Can use diverse cell lines
-
•
High specificity: less off-target effects
-
•
Generates null phenotype
-
•
Viral transduction works better for suspension cells than transfection
-
•
Good format for suspension cells
-
•
Finds receptors, entry factors, and associated genes
-
•
High specificity
-
•
Long-term screens (> 10 days)
-
•
Can inhibit or activate gene expression (CRISPRa and CRISPRi)
-
•
Active in the nucleus
-
•
Can remove large sections of a targeted locus (e.g., inactivate lncRNA genes)
-
•
First-generation reagents graciously shared at low cost on Addgene
-
•
Low cost to perform survival screens
|
| Weaknesses |
-
•
Off-target effects
-
•
False negatives
-
•
Hypomorphs can produce false negatives
-
•
Loss-of-function only
-
•
RISC has questionable or limited activity in the nucleus
-
•
Difficult to transfect primary cells or suspension cells
-
•
Difficult to use suspension cells in an arrayed format
-
•
Expensive to purchase, use, and maintain libraries
-
•
Requires expensive high-throughput microscope or plate reader for analysis
|
-
•
Off-target effects
-
•
False negatives
-
•
PCR/next-gen sequencing needed to identify hits
-
•
Loss-of-function only
-
•
RISC has questionable or limited activity in the nucleus
-
•
Cannot do cell biology or imaging screens
-
•
Target knockdown more difficulty due to only one shRNA-producing provirus per cell
|
-
•
Random insertion mutagenesis cannot specifically target a gene
-
•
Only two available haploid cell lines
-
•
PCR/next-gen sequencing needed to identify hits
-
•
Loss-of-function only
-
•
Retroviral insertion bias may not permit saturation
-
•
Cannot do cell biology or imaging screens
-
•
Arrayed format is subgenomic and requires long-term culturing and storage of many thousands of cell lines with likely high cost
|
-
•
PCR/next-gen sequencing needed to identify hits
-
•
Relatively slower validation
-
•
Cannot do cell biology or imaging screens
-
•
Arrayed lentiviral format will be cumbersome
-
•
Arrayed transfectable CRISPR components (sgRNAs, Thermo, and IDT) are subgenomic at present with whole-genome reagents likely obtained at high cost
|
