Forward and reverse genetics are techniques that can be used to determine the phenotypic and genotypic basis of a given genotype or phenotype respectively.
Forward genetics involves taking a phenotype and attempting to determine the genetic basis of it. After the creation of a mutant library, the bacteria are screened for the phenotype. Any not expressing the phenotype of interest can be sequenced the genes involved identified. An issue with this can be polarity effects. If a mutation is present early in an operon, it can knock out the entire operon. This is not ideal, as it is then difficult to determine which gene is responsible for the phenotype of interest. Repeating this with different mutants can allow the successful identification of the gene, however.
Reverse genetics is the opposite, taking a gene and attempting to determine the phenotype it causes. By searching bioinformatics databases, it is possible to identify whether the gene has been studied before. If the gene has not been studied before, then the gene should be mutated in vivo and then overexpressed. Using RNA-seq, any changes to the transcriptome can be observed, and the resulting phenotype can be identified. To identify the phenotype, phenotypic microarrays (BiOLOG) are used. These are 96-well plates containing different growth media in each well, allowing the identification of phenotypes. However, this approach is trial-and-error based, and may not result in the phenotype being identified. Therefore, this can be costly and futile.
