Construction of A Reference Sequence Data Set For The Human Microbiome Project

人类的生理和健康高度依赖于居住在人体内外多个生态位的数十亿微生物的活动。这些微生物中很少有特征，这在很大程度上是因为大多数还有待培养。因此，这一数量庞大的物种对人类健康的影响仍有待阐明。本提案介绍了J. Craig Venter研究所生成大量参考物种（包括与人体相关的细菌、真菌、病毒和噬菌体）基因组序列的方法。我们还描述了如何从从多个个体的不同身体部位收集的DNA样本中生成宏基因组序列。

Human physiology and health is highly dependent on the activities of billions of microorganisms that inhabit multiple niches within and on the human body. Few of these microorganisms have been characterized, in large part because the majority are yet to be cultivated. As such, the impact this vast number of species has on human health remains to be elucidated. This proposal presents the J. Craig Venter Institute's approach to generating the genomic sequences of a significant number of reference species including bacteria, fungi, viruses and phage that are associated with the human body. We also describe how we will generate metagenomic sequences from DNA samples collected from various body sites of multiple individuals. The specific aims include the sequencing, assembly and annotation of 400 microbial genomes (60 of which will be closed) to provide reference genomes for metagenomic data; characterization of the microbiota from multiple body sites via shotgun sequencing of these samples, and the use of our state-of-the-art assembly and analysis tools to further interpret this data. We will also investigate microeukaryotic diversity, focusing on the 28S rRNA gene and internal transcribed spacer regions. Cell sorting, combined with whole genome amplification will be employed to obtain genomic DNA from previously uncultured prokaryotes. Finally, the dynamic range of expressed RNA from marker genes identified through metagenomic analyses will be interrogated using a novel mRNA transcript capture and amplification approach. This study will be conducted using cutting edge technologies including next generation approaches to sequencing. We will continue outreach and education to research clinicians, and will make all data readily available. Without a doubt, the findings from the proposed study will increase current understanding of the microbial diversity associated with the human body and the impact of these species on health and disease.