Current and future resources for functional metagenomics

Abstract

Functional metagenomics is a powerful experimental approach for studying gene function, starting from the extracted DNA of mixed microbial populations. A functional approach relies on the construction and screening of metagenomic libraries-physical libraries that contain DNA cloned from environmental metagenomes. The information obtained from functional metagenomics can help in future annotations of gene function and serve as a complement to sequence-based metagenomics. In this Perspective, we begin by summarizing the technical challenges of constructing metagenomic libraries and emphasize their value as resources. We then discuss libraries constructed using the popular cloning vector, pCC1FOS, and highlight the strengths and shortcomings of this system, alongside possible strategies to maximize existing pCC1FOS-based libraries by screening in diverse hosts. Finally, we discuss the known bias of libraries constructed from human gut and marine water samples, present results that suggest bias may also occur for soil libraries, and consider factors that bias metagenomic libraries in general. We anticipate that discussion of current resources and limitations will advance tools and technologies for functional metagenomics research.

Keywords: RK2; cloning bias; cosmid library; fosmid library; functional metagenomics; library bias; metagenomic library; pCC1FOS.

Figure 1

Metagenomic libraries exhibit cloning bias when compared to the original environmental sample. (A) Steps involved in the construction of a metagenomic library, from original environmental sample to the final library in the E. coli host (adapted from Lam and Charles, 2015). (B) Relative abundance of bacterial phyla from two previously constructed metagenomic libraries, a human fecal library (Lam and Charles, 2015), and a corn field soil library (Cheng et al., 2014), compared to their original sample DNA extracts. (C) Number of OTUs identified from corn field soil DNA extract and library, and whether the OTUs were present in the library sample only, the extract sample only, or present in both. (D) Examination of cloning bias by comparing the relative abundance of OTUs that were present in both the DNA extract and the cosmid library, shown on a log scale; horizontal line at 1 denotes equal relative abundance in both samples.