Rumen microbiota is a complex biological system, with huge amount of genetic and ecological resource. Metagenomics as “the application of modern genomics techniques to the study of communities of microbial organisms directly in their natural environments, bypassing the need for isolation and lab cultivation of individual species” shows strong vitality in research of the metabolic pathway and exploring the gene resource of rumen microbiota. The application of metagenomics may solve many hot and difficult problems in rumen nutrition. Therefore, the construction and selection procedure were introduced in this paper. The methods of target gene/genome enrichment, the selection principle of vectors, the procedure of nucleic acid extraction, and the application of metagenomics in rumen were related in detail. ［Chinese Journal of Animal Nutrition, 2010，22（3）:527-535］