Interplay of metagenomics and in vitro compartmentalization.
Bottom Line: In recent years, the application of approaches for harvesting DNA from the environment, the so-called, 'metagenomic approaches' has proven to be highly successful for the identification, isolation and generation of novel enzymes.Functional screening for the desired catalytic activity is one of the key steps in mining metagenomic libraries, as it does not rely on sequence homology.In particular, we focus on the use of in vitro compartmentalization (IVC) approaches to address potential advantages and problems the merger of culture-independent and IVC techniques might bring on the mining of enzyme activities in microbial communities.
Affiliation: CSIC, Institute of Catalysis, Department of Applied Biocatalysis, Madrid, Spain. firstname.lastname@example.orgShow MeSH
Related in: MedlinePlus
Mentions: In vitro compartmentalization is based on water‐in‐oil emulsions, where the water phase is dispersed in the oil phase to form microscopic aqueous compartments. Each droplet contains, on average, a single gene, and serves as an artificial cell allowing for transcription, translation and the activity of the resulting proteins, to take place within the compartment. The oil phase remains largely inert and restricts the diffusion of genes and proteins between compartments (Fig. 3). The droplet volume (∼5 femtoliter) enables a single DNA molecule to be transcribed and translated (Griffiths and Tawfik, 2006), as well as the detection of single enzyme molecules (Griffiths and Tawfik, 2003). The high capacity of the system (> 1010 in 1 ml of emulsion), the ease of preparing emulsions and their high stability over a broad range of temperatures, render IVC an attractive system for HTS of enzymes, as well as for many other HT genetic and genomic manipulations (for a recent review, see Griffiths and Tawfik, 2006).
Affiliation: CSIC, Institute of Catalysis, Department of Applied Biocatalysis, Madrid, Spain. email@example.com