In order to understand how information about epistatic interactions relates to gene pathways, consider a simple example of vulval cell differentiation in C. elegans.
In both cases, the resulting phenotype is marked by an absence of vulval cells as there is an upstream block in the differentiation pathway.
Both SGA and dSLAM approaches rely on these yeast knockout strains which are transformed/mated to generate haploid double mutants.
In order to develop a richer understanding of genetic interactions, experimental approaches are shifting away from this binary classification of phenotypes as wild type or synthetic lethal.
The E-MAP approach is particularly compelling because of its ability to highlight both alleviating and aggravating effects and this capacity is what distinguishes this method from others such as SGA and dSLAM.
Clusters organized by sub-cellular localization and general cellular processes (e.g. cell cycle) have yielded profitable results in S. cerevisiae.
Collins et al. (2007) carried out a comparison of E-MAP scores and physical interaction data from large-scale affinity purification methods (AP-MS) and their data demonstrate that an E-MAP approach identifies protein-protein interactions with a specificity equal to that of traditional methods such as AP-MS .
High throughput methods of examining epistatic relationships face difficulties, however as the number of possible gene pairs is extremely large (~20 million in S. cerevisiae) and the estimated density of genetic interactions is quite low.
The generation of libraries of essential gene mutants presents significant difficulties however, as these mutations have a lethal phenotype.
The decreased abundance by messenger RNA perturbation (DAmP) strategy is particularly common for the high-throughput generation of mutants necessary for this kind of analysis and allows for the partial disruption of essential genes without loss of viability.
[9] DAmP relies upon the destabilization of mRNA transcripts by integrating an antibiotic selectable marker into the 3’UTR, downstream of the stop codon (figure 2).
mRNA’s with 3’ extended transcripts are rapidly targeted for degradation and the result is a downregulation of the gene of interest while it remains under the control of its native promoter.
Tagging at the deletion sites with molecular barcodes, unique 20-bp sequences, allows for the identification and study of relative fitness levels in each mutant strain.