Considerations and best practices in animal science 16S ribosomal RNA gene sequencing microbiome studies.
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| Title: | Considerations and best practices in animal science 16S ribosomal RNA gene sequencing microbiome studies. |
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| Authors: | Weinroth, Margaret D1 (AUTHOR) maggie.weinroth@usda.gov, Belk, Aeriel D2,3 (AUTHOR), Dean, Chris4 (AUTHOR), Noyes, Noelle4 (AUTHOR), Dittoe, Dana K5 (AUTHOR), Rothrock, Michael J1 (AUTHOR), Ricke, Steven C5 (AUTHOR), Myer, Phillip R6 (AUTHOR), Henniger, Madison T6 (AUTHOR), Ramírez, Gustavo A7 (AUTHOR), Oakley, Brian B7 (AUTHOR), Summers, Katie Lynn8 (AUTHOR), Miles, Asha M8 (AUTHOR), Ault-Seay, Taylor B6 (AUTHOR), Yu, Zhongtang9 (AUTHOR), Metcalf, Jessica L2 (AUTHOR), Wells, James E10 (AUTHOR) |
| Source: | Journal of Animal Science. Feb2022, Vol. 100 Issue 2, p1-18. 18p. |
| Document Type: | Article |
| Subjects: | Animal science, Ribosomal RNA, Molecular biology, Best practices, Microbial ecology |
| Abstract: | Microbiome studies in animal science using 16S rRNA gene sequencing have become increasingly common in recent years as sequencing costs continue to fall and bioinformatic tools become more powerful and user-friendly. The combination of molecular biology, microbiology, microbial ecology, computer science, and bioinformatics—in addition to the traditional considerations when conducting an animal science study—makes microbiome studies sometimes intimidating due to the intersection of different fields. The objective of this review is to serve as a jumping-off point for those animal scientists less familiar with 16S rRNA gene sequencing and analyses and to bring up common issues and concerns that arise when planning an animal microbiome study from design through analysis. This review includes an overview of 16S rRNA gene sequencing, its advantages, and its limitations; experimental design considerations such as study design, sample size, sample pooling, and sample locations; wet lab considerations such as field handing, microbial cell lysis, low biomass samples, library preparation, and sequencing controls; and computational considerations such as identification of contamination, accounting for uneven sequencing depth, constructing diversity metrics, assigning taxonomy, differential abundance testing, and, finally, data availability. In addition to general considerations, we highlight some special considerations by species and sample type. [ABSTRACT FROM AUTHOR] |
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| Author Affiliations: | 1U.S. Department of Agriculture, Agricultural Research Service, U.S. National Poultry Research Center (USNPRC) , Athens, GA 30605 , USA 2Department of Animal Sciences, Colorado State University , Fort Collins, CO 80524 , USA 3Joint Institute of Food Safety and Applied Nutrition, University of Maryland , College Park, MD 20740 , USA 4Department of Veterinary Population Medicine, University of Minnesota , St. Paul, MN 55108 , USA 5Meat Science and Animal Biologics Discovery Program, Department of Animal and Dairy Sciences, University of Wisconsin , Madison, WI 53706 , USA 6Department of Animal Science, University of Tennessee , Knoxville, TN 37996 , USA 7College of Veterinary Medicine, Western University of Health Sciences , Pomona, CA 91766 , USA 8U.S. Department of Agriculture, Agricultural Research Service, Beltsville Agricultural Research Center (BARC) , Beltsville, MD 20705 , USA 9Department of Animal Sciences, The Ohio State University , Columbus, OH 43210 , USA 10USDA ARS US Meat Animal Research Center (USMARC) , Clay Center, NE 68933 , USA |
| Full Text Word Count: | 17583 |
| ISSN: | 0021-8812 |
| DOI: | 10.1093/jas/skab346 |
| Accession Number: | 155109339 |
| Database: | Veterinary Source |
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| Abstract: | Microbiome studies in animal science using 16S rRNA gene sequencing have become increasingly common in recent years as sequencing costs continue to fall and bioinformatic tools become more powerful and user-friendly. The combination of molecular biology, microbiology, microbial ecology, computer science, and bioinformatics—in addition to the traditional considerations when conducting an animal science study—makes microbiome studies sometimes intimidating due to the intersection of different fields. The objective of this review is to serve as a jumping-off point for those animal scientists less familiar with 16S rRNA gene sequencing and analyses and to bring up common issues and concerns that arise when planning an animal microbiome study from design through analysis. This review includes an overview of 16S rRNA gene sequencing, its advantages, and its limitations; experimental design considerations such as study design, sample size, sample pooling, and sample locations; wet lab considerations such as field handing, microbial cell lysis, low biomass samples, library preparation, and sequencing controls; and computational considerations such as identification of contamination, accounting for uneven sequencing depth, constructing diversity metrics, assigning taxonomy, differential abundance testing, and, finally, data availability. In addition to general considerations, we highlight some special considerations by species and sample type. [ABSTRACT FROM AUTHOR] |
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| ISSN: | 00218812 |
| DOI: | 10.1093/jas/skab346 |