Investigating the Role of let-7 in Rotifer Aging

The recent discovery of microRNAs (miRNAs) in Caernohabditis elegans has ignited researchers to study its role in gene regulation. These small (~22 nucleotides), non-coding RNAs have been implicated in post-transcriptional regulation of protein-coding mRNAs. Together with lin-4, let-7 was initially discovered in 2000. Now, homologues of this miRNA have been identified in most animals. This miRNA is reported to regulate the expression of aging-related mRNA daf-12 in C. elegans. Our intent is to correlate the expression of let-7 to the aging process of the monogonont Brachionous manjavacas. This brackish-water rotifer is characteristic for having both sexual and asexual reproduction. Females that produce eggs by mitosis are considered to be amictic, and their eggs produce other amictic females. However, overcrowding signals amictic females to hatch mictic females, whose eggs carry haploid males. This species’ growing genomic and transcriptomic information allows researchers to use it as a model system. In addition, its three-week lifespan and manageable size (~ 1mm, 1000 nuclei) make it an excellent model for aging research. RNA will be extracted at the four stages of the amictic female’s life (egg, neonate, reproductive and post-reproductive); which will be reverse transcribed into cDNA.  Amplifying with qPCR, the levels of expression will be quantified. A synthetic let-7 RNA oligo will be used as a control to establish its presence in these animals. Preliminary results have demonstrated that we can obtain sufficient amplification of let-7 cDNA from as few as ten animals.