Microbial diversity analysis of water samples from various hydrocarbon resource environments using next-generation 16S pyrosequencing
Microbial diversity analysis of water samples from various hydrocarbon resource environments using next-generation 16S pyrosequencing
Water samples from different oil and gas company sites were sampled and sent with the objective of microbial community analysis using 16S pyrotag sequencing. Sample was collected from various hydrocarbon resource environmental sites and consisted of high temperature water sample, high salinity water sample, sour-process water sample, and salt water disposal tank water samples. All the samples were collected from the surface and sampling was performed by grab sampling (all of the test material was collected at one time). Water samples were passed through a 0.2 µm filter with the collected biomass stored at -70°C. DNAs were isolated using the modified Marmur’s protocol with bead-beating. DNAs from solid samples (e.g. cores or oil sands) were isolated with a FastDNA Spin Kit for Soil. The 16S rDNA genes from the isolated community DNAs were PCR-amplified using universal primers. Bar-codes were then added to the PCR products (representing the resident communities) using a minimal number of PCR cycles with bar-coded primers. Following pyrosequencing, the 16S rDNA sequences determined were used to derive the composition of the microbial communities. This research was supported by funding from Genome Canada, Genome Alberta, the Government of Alberta and Genome BC.
Investigations of the microbial communities present in oil and gas reserves have traditionally relied on culture based techniques. Traditional culture based approaches to environmental microbiology can miss up to 90% of the indigenous community, only culture-independent approaches (metagenomics) can capture the true indigenous microbial diversity. The objective of our study is to generate, develop, deploy and store metagenomic database of the microbial communities found in hydrocarbon resource environments. In order to achieve this, next-generation pyrosequencing technologies were used for a 16S rRNA gene-based profiling of microbial communities in a variety of hydrocarbon resource environments.