Evaluation of in-field stability of mitochondrial and nuclear DNA in snow sampled fecal pellets from Rock ptarmigan (Lagopus muta)

Abstract

Non-invasive genetic sampling (NGS) is highly relevant in genetic studies applied to ecology. Methods should be tailored to species and environments. In this context, the in-field stability of fecal DNA is essential. Here we describe a method to test the stability of DNA in fecal pellets (FPs) of rock ptarmigan (Lagopus muta). Rock ptarmigan spend their entire lifecycle in the alpine region and roost in individual snow holes to save energy and avoid predation. Roosting birds commonly deposit FPs and individual snow roosts are not reused. FPs (n=146) were collected from 20 individual rock ptarmigan snow roosts in the Lifjell Mountains in Telemark County, southeast Norway between 17 January and 7 July, 2010. DNA was extracted from the 146 samples and we tested in-field temporal stability in mitochondrial control region DNA, nuclear DNA, chromo-helicase-DNA binding (CHD) gene and three microsatellites (msats). MtDNA and CHD gene were scored on polyacrylamide gel while msats were sized by capillary electrophoresis. For mtDNA, the mean stability in 20 roosts was 74 and 59 days respectively in a best and worst case scenario. For the CHD gene, the mean stability was 122 and 88 days and for the 3 msats, mean stability varied between 39 - 58 and 21 - 46 days respectively in the two scenarios. The CHD gene had the highest stability. MtDNA had significantly higher stability than msats and between individual msats stability varied significantly. DNA degradation appeared to accelerate considerably when frozen, snow-covered FPs eventually became exposed to higher temperatures and precipitation during the spring snow-melt. FPs from rock ptarmigan snow roosts appear to be a reliable DNA source when analyzing mtDNA (species determination) and nDNA (sex determination and msat analysis). These methods may also be applicable to other snow-roosting grouse.