Antibiotics pp 307-329 | Cite as

Functional Metagenomics to Study Antibiotic Resistance

Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1520)

Abstract

The construction and screening of metagenomic expression libraries has great potential to identify novel genes and their functions. Here, we describe metagenomic library preparation from fecal DNA, screening of libraries for antibiotic resistance genes (ARGs), massively parallel DNA sequencing of the enriched DNA fragments, and a computational pipeline for high-throughput assembly and annotation of functionally selected DNA.

Key words

Functional metagenomics Antibiotic resistance genes Resistome Functional selections Massively parallel DNA sequencing High-throughput assembly Profile HMM-based annotation PARFuMS Resfams 

Notes

Acknowledgments

The original implementation of functional metagenomics selections for interrogating resistance genes was described by J. Handelsman and colleagues in 1998. We thank A. Moore and B. Wang for protocol optimization of high-throughput versions of this method presented in this manuscript, A. Reyes and K. Forsberg for computational implementation of PARFuMS, M. Gibson for development of the Resfams database, and members of the Dantas lab for helpful general discussions and for comments on the manuscript.

This work was supported in part by the NIH Director’s New Innovator Award (http://commonfund.nih.gov/newinnovator/), the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK:http://www.niddk.nih.gov/), and the National Institute of General Medical Sciences (NIGMS: http://www.nigms.nih.gov/), of the National Institutes of Health (NIH) under award numbers DP2DK098089 and R01GM099538 to G.D. The content is solely the responsibility of the authors and does not necessarily represent the official views of the funding agencies.

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Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • Manish Boolchandani
    • 1
  • Sanket Patel
    • 1
    • 2
  • Gautam Dantas
    • 1
    • 2
    • 3
    • 4
  1. 1.Center for Genome Sciences and Systems BiologyWashington University School of MedicineSt. LouisUSA
  2. 2.Department of Pathology and ImmunologyWashington University School of MedicineSt. LouisUSA
  3. 3.Department of Biomedical EngineeringWashington UniversitySt. LouisUSA
  4. 4.Department of Molecular MicrobiologyWashington University School of MedicineSt. LouisUSA

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