IMIS

Samples were collected by a diver by cutting squares of mat from the lake floor and gently lifting them into a plastic box previously cleaned with antibacterial wipes. Box lids were sealed under water and samples returned to the surface and transferred to lakeside laboratory, where they were immediately dissected using flame-sterilized blades and forceps. Pinnacle and ridge-pit mats were dissected according to their distinctively pigmented horizontal upper (pink-purple and brown-purple, respectively), middle (pale purple and green-beige, respectively), and lower (all beige) layers. Prostrate mats were similarly dissected into upper (brown-purple), middle (green- beige), and lower (beige) layers. Dissected subsamples were rinsed in ster- ile deionized water, transferred into sterile plastic tubes, and frozen at 20°C until further analysis.
Study Extent: Microbial mat samples representing the main three macroscopic mat morphologies, i.e., cuspate pinnacle, ridge-pit, and prostrate were collected from the floor of Lake Fryxell in November 2012.
Method step description:

1. DNA extractions of each of the triplicate mat sections were performed using the MoBio PowerDNA biofilm kit according to the manufacturer’s instructions. DNA was quantified using a Qubit fluorometer, and equal amounts of DNA per sample were pooled for 16S rRNA gene amplification. 16S rRNA gene PCR products were amplified in triplicate, pooled, and cleaned using the MoBio UltraClean PCR clean-up kit ac- cording to the manufacturer’s instructions. For the high-throughput se- quencing, we used primers (515f and 806r), described by Caporaso et al., which amplify both archaeal and bacterial 16S rRNA genes, including cyanobacterial 16S rRNA genes.
2. Sequencing was performed on a MiSeq Illumina sequencer at NZ Genomics Limited, Palmerston North, New Zealand.