112 -- Purifying Total RNA to Explore the Role of piRNAs in Zebrafish Stress Response

Tuna fish plays a significant economic and environmental role and contributes at least $40 billion annually to the global economy, but climate change has been rendering tuna populations particularly vulnerable to rising ocean temperatures and acidification. Recent studies have shown the emerging role of noncoding RNA (ncRNAs) such as piRNA in stress response and transgenerational inheritance. In order to assess the potential role of piRNA in tuna population’s response to climate stress, we use the zebrafish as a laboratory model to identify piRNAs whose steady-state levels change in response to conditions mimicking global climate change stress: elevated temperature, decreased pH and reduced nutrient availability. Additionally, single nucleotide polymorphisms (SNPs) in piRNA could be used as biological markers for predicting and determining which populations of zebrafish and tuna are under environmental stress. In order to understand how specific piRNAs are involved in the stress response, we will accomplish our goal through a two-pronged approach. The first approach will be focused on using pre-selected candidate piRNA to test for stress-response. The second approach will analyze the genome-wide transcriptome set of total piRNA in zebrafish samples using NERD-seq, a method that allows us to analyze piRNA more effectively at the genome-wide scale. Here we report on our effort to purify total RNA from selected tissues of stressed and unstressed zebrafish for use in quantifying candidate piRNA levels by RT-qPCR method.