Analysis of unc-22 mutant phenotype in Caenorhabditis elegans, Odey, Ochuko; Villanueva, Grace
College of Science and Mathematics
Caenorhabditis elegan’s unc-22 codes for UNC-22 protein (twitchin), a homolog of proteins MyBP-C (cardiac myosin binding protein) and titin in humans. The unc-22 gene was identified as the founding member of a family of very large cytoplasmic proteins, on which muscle structure and function depends. MyBP-C is found to be frequently mutated in cardiomyopathy patients, while mutations in titin are associated with loss of muscle function which is one of the characteristics of muscular dystrophy. The knockdown of unc-22 was initiated by RNA interference (RNAi). C. elegans are the ideal specimens when working with RNAi due to the many effective techniques in which they can be introduced to specific double stranded RNA by feeding them bacteria. In this experiment, we fed the nematodes bacteria expressing dsRNA of unc-22 gene to trigger the silencing of the protein (twitchin). Knockdown of this gene is expected to produce a twitching muscular movement phenotype. In this case, we used IPTG and Carbenicillin in nematode growth agar (NGM) to induce silencing of the unc-22 gene and select for antibiotic resistant E.coli respectively. Varying the concentration of IPTG we will observe the number of twitches, and the distance traveled per/min of mutant worms growing in the different concentrations. We will then synchronize the worms by bleaching in order to place them in the same stage of development, and allow us to plate the eggs on HT115/NC and HT115/NIC in order to compare their hatching rates. In a follow up experiment, we will also place eggs from L2 stage worms on new growth media plates containing HT115 E. coli/NC and observe if gene silencing will continue to the F1 generation. The number of twitches/min in the F1 generation will be recorded. Results will be presented at the symposium.
