How will coldwater fish survive on a hot planet?

Methods

Summary

The laboratory work for this project will occur in a few major stages. Starting with tissue samples collected from brook trout across the species range, I will begin by extracting and quantifying the DNA. Once the DNA has been extracted from the tissue and verified for quality and quantity I will begin library preparation, which is a process that prepares the DNA for genetic sequencing. Following library preparation, I will use a bait capture approach to selectively target regions of the genome for sequencing using the custom panel we designed in concert with Arbor Biosciences. Following the bait capture reaction, we will perform genetic sequencing on the samples using next generation sequencing. 

Challenges

Since this is the first time that these baits have been designed and are being tested, I suspect there were be significant challenges associated with the lab work for this project. There is the possibility that there may be baits that fail during the actual reaction, that otherwise would have survived the simulated quality control conditions during the design phase. There is also the possibility of off-target sequences being captured by our custom baits. I think that generating this data may require some expertise with laboratory techniques that are nascent to the field of genomics; however, I am confident that I have the skills and the resources needed to be successful in completing this lab work.

Pre Analysis Plan

I plan to prepare and analyzed these data using a standard bioinformatics pipeline for capture sequencing. While there is not currently a reference genome available for Salvelinus fontinalis, there is a reference genome for a sister species, the Dolly Varden, Salvelinus malma. Following processing of the sequencing data, I will perform quality control and call single nucleotide polymorphisms (SNPs). I will then analyze the SNPs and examine the genetic distinction among individuals and populations from differing environments across the species range.