Department of Medical Microbiology and Infection Control, VU University Medical Center, Amsterdam, The Netherlands.
FASEB J. 2010 Nov;24(11):4556-64. doi: 10.1096/fj.10-156190. Epub 2010 Jul 19.
The human intestinal microbiota is known to play an important role in human health and disease, and with the advent of novel molecular techniques, disease-specific variations in its composition have been found. However, analysis of the intestinal microbiota has not yet been applicable in large-scale clinical research or routine diagnostics because of the complex and expensive nature of the techniques needed. Here, we describe a new PCR-based profiling technique for high-throughput analysis of the human intestinal microbiota, which we have termed IS-pro. This technique combines bacterial species differentiation by the length of the 16S-23S rDNA interspace region with instant taxonomic classification by phylum-specific fluorescent labeling of PCR primers. We validated IS-pro in silico, in vitro, and in vivo, on human colonic biopsies and feces, and introduced a standardized protocol for data analysis. IS-pro is easy to implement in general clinical microbiological laboratories with access to capillary gel electrophoresis, and the high-throughput nature of the test makes analysis of large numbers of samples feasible. This combination renders IS-pro ideally suited for use in clinical research and routine diagnostics.
人类肠道微生物群被认为在人类健康和疾病中发挥着重要作用，随着新型分子技术的出现，人们发现其组成在疾病方面存在特异性变化。然而，由于所需技术的复杂性和昂贵性质，肠道微生物组的分析尚未在大规模临床研究或常规诊断中得到应用。在这里，我们描述了一种新的基于 PCR 的高通量分析人类肠道微生物组的分析技术，我们称之为 IS-pro。该技术通过 16S-23S rDNA 间隔区的长度来区分细菌种类，并通过针对 PCR 引物的属特异性荧光标记进行即时分类。我们在计算机模拟、体外和体内对 IS-pro 进行了验证，涉及人类结肠活检和粪便，并引入了标准化数据分析方案。IS-pro 易于在一般临床微生物学实验室中实施，这些实验室可以使用毛细管凝胶电泳，并且该测试的高通量性质使得分析大量样本成为可能。这种组合使 IS-pro 非常适合用于临床研究和常规诊断。
