Rooks David J, McDonald James E, McCarthy Alan J
Institute of Integrative Biology, University of Liverpool, Liverpool, United Kingdom.
Methods Enzymol. 2012;510:375-94. doi: 10.1016/B978-0-12-415931-0.00020-3.
Most of the microorganisms responsible for nutrient cycling in the environment have yet to be cultivated, and this could include those species responsible for the degradation of cellulose. Known cellulases are well defined at the protein sequence level, but gene variants are difficult to amplify from environmental DNA. The identification of novel cellulase genes independent of DNA amplification is made possible by adopting a direct metagenome sequencing approach to provide genes that can be cloned, expressed, and characterized prior to potential exploitation, all in the absence of any information on the species from which they originated. In this chapter, emerging strategies and methods that will enable the identification of novel cellulase genes and provide an unbiased perspective on gene expression in situ are presented.
环境中负责养分循环的大多数微生物尚未得到培养，这可能包括那些负责纤维素降解的物种。已知的纤维素酶在蛋白质序列水平上已得到很好的定义，但基因变体很难从环境DNA中扩增出来。通过采用直接宏基因组测序方法，可以在不依赖DNA扩增的情况下鉴定新的纤维素酶基因，从而提供可在潜在开发之前进行克隆、表达和表征的基因，而无需任何关于其来源物种的信息。在本章中，将介绍能够鉴定新的纤维素酶基因并提供原位基因表达无偏视角的新兴策略和方法。
