N-168. Metagenomic and Single-Cell Genomics of Greenland Glacial Ice

J. M. Gosses, B. D. Lanoil, S. Han;
Univ. of California, Riverside, CA.

A variety of recent studies of ice cores indicates the presence of a microbial assemblage that may be active and adapted to life in the ice. To understand the mechanisms of adaptation to the extreme cold, dark and isolated (hence oligotrophic), and potentially high salt and/or low pH, we have undertaken a metagenomic approach. Due to the extremely low abundance of cells and paucity of sample, we developed an extensive hypochlorite bleach-based decontamination protocol and DNA cell concentration and extraction protocols. As a test of our protocols, we decontaminated, melted, and concentrated particles from a sample of ice from the GISP2 core. After decontamination and melting the cell count using DAPI was determined to be 3.2 x 103 cells ml-1 and the sample was concentrated 343 fold. DNA was extracted using the Zygem enzyme, and multistrand displacement amplification (MDA) with the phi29 DNA polymerase provided sufficient genomic DNA for PCR amplification. DGGE analysis of 16S rDNA showed low diversity, with bands related to Acinetobacter, Capnocytophaga, Enterobacter, Sphingopyxis, Propionibacterium, Lactobacillus and Ralstonia. We are currently characterizing functional genes from this sample as well. Single cells were also sorted and counted by flow cytometry from the sample and genomic DNA was obtained via MDA. Initial results show similar 16S rRNA gene sequences in these single cell genomes as the metagenomic product. These protocols are broadly useful for all ice core samples, and the resulting metagenomic and single cell genomic sequences will provide insights into adaptations to life in ice.