Development of a Genome-Proxy Microarray for Profiling Marine Microbial Communities and its Application to a Time Series in Monterey Bay, California

摘要

This thesis describes the development and application of a new tool for profiling marine microbial communities. Chapter 1 places the tool in the context of the range of methods used currently. Chapter 2 describes the development and validation of the 'genome proxy' microarray, which targeted marine microbial genomes and genome fragments using sets of 70-mer oligonucleotide probes. In a natural community background, array signal was highly linearly correlated to target cell abundance (R2 of 1.0), with a dynamic range from 10-10 cells/mI. Genotypes with >--80% average nucleotide identity to those targeted cross-hybridized to target probesets but produced distinct, diagnostic patterns of hybridization. Chapter 3 describes the development an expanded array, targeting 268 microbial genotypes, and its use in profiling 57 samples from Monterey Bay. Comparison of array and pyrosequence data for three samples showed a strong linear correlation between target abundance using the two methods (R2=0.85-0.91). Array profiles clustered into shallow versus deep, and the majority of targets showed depth-specific distributions consistent with previous observations. Although no correlation was observed to oceanographic season, bloom signatures were evident. Array-based insights into population structure suggested the existence of ecotypes among uncultured clades. Chapter 4 summarizes the work and discusses future directions.