Petrone Joseph R, Muñoz-Beristain Alam, Glusberger Paula Rios, Russell Jordan T, Triplett Eric W
Microbiology and Cell Science Department, Institute of Food and Agricultural Sciences, University of Florida, Gainesville, FL 32603, USA.
Microorganisms. 2022 Feb 26;10(3):513. doi: 10.3390/microorganisms10030513.
With the current advancements in DNA sequencing technology, the limiting factor in long-read metagenomic assemblies is now the quantity and quality of input DNA. Although these requirements can be met through the use of axenic bacterial cultures or large amounts of biological material, insect systems that contain unculturable bacteria or that contain a low amount of available DNA cannot fully utilize the benefits of third-generation sequencing. The citrus greening disease insect vector is an example that exhibits both of these limitations. Although endosymbiont genomes have mostly been closed after the short-read sequencing of amplified template DNA, creating de novo long-read genomes from the unamplified DNA of an insect population may benefit communities using bioinformatics to study insect pathosystems. Here all four genomes of the infected microbiome were sequenced to closure using unamplified template DNA and two long-read sequencing technologies. Avoiding amplification bias and using long reads to assemble the bacterial genomes allowed for the circularization of the endosymbiont of for the first time and paralleled the annotation context of all four reference genomes without utilizing a traditional hybrid assembly. The strategies detailed here are suitable for the sequencing of other insect systems for which the input DNA, time, and cost are an issue.
随着DNA测序技术的当前进展，长读长宏基因组组装中的限制因素现在是输入DNA的数量和质量。尽管可以通过使用无菌细菌培养物或大量生物材料来满足这些要求，但含有不可培养细菌或可用DNA量低的昆虫系统无法充分利用第三代测序的优势。柑橘黄龙病昆虫载体就是一个同时存在这两种限制的例子。尽管在扩增模板DNA的短读长测序后，内共生体基因组大多已完成封闭，但从昆虫群体的未扩增DNA创建从头长读长基因组可能会使利用生物信息学研究昆虫病理系统的群体受益。在这里，使用未扩增的模板DNA和两种长读长测序技术对受感染微生物组的所有四个基因组进行了测序以实现封闭。避免扩增偏差并使用长读长来组装细菌基因组首次实现了对的内共生体的环化，并在不使用传统混合组装的情况下与所有四个参考基因组的注释背景平行。这里详细介绍的策略适用于输入DNA、时间和成本是问题的其他昆虫系统的测序。
