Submission Title

Validation of a Requirement for Regena/NOT-2 in miRNA Mediated Gene Silencing

Location

Jereld R. Nicholson Library

Subject Area

Biology

Description

miRNAs are small non-coding RNAs that silence gene expression. A forward genetic screen led to the discovery of a mutant with defective gene silencing that contains a mutation in the gene Regena/NOT2. We hypothesize that the defect in silencing is a result of the mutation in Regena/NOT2 rather than another mutation inadvertently generated in the genetic screen. To formally test this hypothesis, we must either perform a genetic rescue, by adding a functional copy of the Regena/NOT2 gene into the mutant fly and observing restoration of silencing or recreating the mutant phenotype by generating an independent mutation in Regena/NOT2 and observing the silencing defect as seen in the original mutant flies. We opted to generate an independent mutation in Regena/NOT2 and assay silencing with a GFP-based reporter of silencing. To do this, we generated flies that contained the new mutation in Regena/NOT2, the reporter of silencing, and the ability to compare silencing in adjacent cells with or without the new mutation in Regena/NOT2. We hypothesize that the independent line of flies containing the new mutation in Regena/NOT2 will show the same defect in silencing as our original mutants.

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Validation of a Requirement for Regena/NOT-2 in miRNA Mediated Gene Silencing

Jereld R. Nicholson Library

miRNAs are small non-coding RNAs that silence gene expression. A forward genetic screen led to the discovery of a mutant with defective gene silencing that contains a mutation in the gene Regena/NOT2. We hypothesize that the defect in silencing is a result of the mutation in Regena/NOT2 rather than another mutation inadvertently generated in the genetic screen. To formally test this hypothesis, we must either perform a genetic rescue, by adding a functional copy of the Regena/NOT2 gene into the mutant fly and observing restoration of silencing or recreating the mutant phenotype by generating an independent mutation in Regena/NOT2 and observing the silencing defect as seen in the original mutant flies. We opted to generate an independent mutation in Regena/NOT2 and assay silencing with a GFP-based reporter of silencing. To do this, we generated flies that contained the new mutation in Regena/NOT2, the reporter of silencing, and the ability to compare silencing in adjacent cells with or without the new mutation in Regena/NOT2. We hypothesize that the independent line of flies containing the new mutation in Regena/NOT2 will show the same defect in silencing as our original mutants.