Variations in dissolved O2 in a Chinese lake drive changes in microbial communities and impact sedimentary GDGT distributions

Next-generation sequencing technologies generate very large numbers of short reads. Even with very deep genome coverage, short read lengths cause problems in de novo assemblies. The use of paired-end libraries with a fragment size shorter than twice the read length provides an opportunity to generate much longer reads by overlapping and merging read pairs before assembling a genome. We present FLASH, a fast computational tool to extend the length of short reads by overlapping paired-end reads from fragment libraries that are sufficiently short. We tested the correctness of the tool on one million simulated read pairs, and we then applied it as a pre-processor for genome assemblies of Illumina reads from the bacterium Staphylococcus aureus and human chromosome 14. FLASH correctly extended and merged reads >99% of the time on simulated reads with an error rate of <1%. With adequately set parameters, FLASH correctly merged reads over 90% of the time even when the reads contained up to 5% errors. When FLASH was used to extend reads prior to assembly, the resulting assemblies had substantially greater N50 lengths for both contigs and scaffolds. The FLASH system is implemented in C and is freely available as open-source code at http://www.cbcb.umd.edu/software/flash. t.magoc@gmail.com.

The ongoing revolution in high-throughput sequencing continues to democratize the ability of small groups of investigators to map the microbial component of the biosphere. In particular, the coevolution of new sequencing platforms and new software tools allows data acquisition and analysis on an unprecedented scale. Here we report the next stage in this coevolutionary arms race, using the Illumina GAIIx platform to sequence a diverse array of 25 environmental samples and three known "mock communities" at a depth averaging 3.1 million reads per sample. We demonstrate excellent consistency in taxonomic recovery and recapture diversity patterns that were previously reported on the basis of metaanalysis of many studies from the literature (notably, the saline/nonsaline split in environmental samples and the split between host-associated and free-living communities). We also demonstrate that 2,000 Illumina single-end reads are sufficient to recapture the same relationships among samples that we observe with the full dataset. The results thus open up the possibility of conducting large-scale studies analyzing thousands of samples simultaneously to survey microbial communities at an unprecedented spatial and temporal resolution.