Liu Guohong, Weston Christopher Q, Pham Long K, Waltz Shannon, Barnes Helen, King Paula, Sphar Dan, Yamamoto Robert T, Forsyth R Allyn
FLIR Systems, Inc., La Jolla, California, 92037, United States of America.
Singlera Genomics, Inc., La Jolla, California, 92037, United States of America.
PLoS One. 2016 Jan 4;11(1):e0146064. doi: 10.1371/journal.pone.0146064. eCollection 2016.
We describe continuing work to develop restriction endonucleases as tools to enrich targeted genomes of interest from diverse populations. Two approaches were developed in parallel to segregate genomic DNA based on cytosine methylation. First, the methyl-sensitive endonuclease HpaII was used to bind non-CG methylated DNA. Second, a truncated fragment of McrB was used to bind CpG methylated DNA. Enrichment levels of microbial genomes can exceed 100-fold with HpaII allowing improved genomic detection and coverage of otherwise trace microbial genomes from sputum. Additionally, we observe interesting enrichment results that correlate with the methylation states not only of bacteria, but of fungi, viruses, a protist and plants. The methods presented here offer promise for testing biological samples for pathogens and global analysis of population methylomes.
我们描述了持续开展的工作，即开发限制性核酸内切酶作为从不同群体中富集目标感兴趣基因组的工具。并行开发了两种基于胞嘧啶甲基化来分离基因组DNA的方法。第一，甲基敏感内切酶HpaII用于结合非CG甲基化DNA。第二，McrB的截短片段用于结合CpG甲基化DNA。使用HpaII时，微生物基因组的富集水平可超过100倍，从而改善了基因组检测，并能覆盖痰液中原本痕量的微生物基因组。此外，我们观察到有趣的富集结果，这些结果不仅与细菌的甲基化状态相关，还与真菌、病毒、原生生物和植物的甲基化状态相关。本文介绍的方法为检测生物样本中的病原体以及群体甲基化组的全局分析带来了希望。
