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Collection and Processing of Lymph Nodes from Large Animals for RNA Analysis: Preparing for Lymph Node Transcriptomic Studies of Large Animal Species
- Source :
- J Vis Exp
- Publication Year :
- 2018
- Publisher :
- MyJove Corporation, 2018.
-
Abstract
- Large animals (both livestock and wildlife) serve as important reservoirs of zoonotic pathogens, including Brucella, Mycobacterium bovis, Salmonella, and E. coli, and are useful for the study of pathogenesis and/or spread of the bacteria in natural hosts. With the key function of lymph nodes in the host immune response, lymph node tissues serve as a potential source of RNA for downstream transcriptomic analyses, in order to assess the temporal changes in gene expression in cells over the course of an infection. This article presents an overview of the process of lymph node collection, tissue sampling, and downstream RNA processing in livestock, using cattle (Bos taurus) as a model, with additional examples provided from the American bison (Bison bison). The protocol includes information about the location, identification, and removal of lymph nodes from multiple key sites in the body. Additionally, a biopsy sampling methodology is presented that allows for a consistency of sampling across multiple animals. Several considerations for sample preservation are discussed, including the generation of RNA suitable for downstream methodologies like RNA-sequencing and RT-PCR. Due to the long delays inherent in large animal vs. mouse time course studies, representative results from bison and bovine lymph node tissues are presented to describe the time course of the degradation in this tissue type, in the context of a review of previous methodological work on RNA degradation in other tissues. Overall, this protocol will be useful to both veterinary researchers beginning transcriptome projects on large animal samples and to molecular biologists interested in learning techniques for in vivo tissue sampling and in vitro processing.
- Subjects :
- 0301 basic medicine
General Chemical Engineering
Context (language use)
Animals, Wild
Brucella
Computational biology
General Biochemistry, Genetics and Molecular Biology
Bison bison
Transcriptome
03 medical and health sciences
Gene expression
medicine
symbols.heraldic_charge
Animals
Lymph node
Immunology and Infection
biology
Bison
General Immunology and Microbiology
General Neuroscience
RNA
biology.organism_classification
030104 developmental biology
medicine.anatomical_structure
symbols
Cattle
Lymph
Lymph Nodes
Subjects
Details
- ISSN :
- 1940087X
- Database :
- OpenAIRE
- Journal :
- Journal of Visualized Experiments
- Accession number :
- edsair.doi.dedup.....92205de30f315389888dd9eb92e5233f