High-molecular-weight DNA extraction for long-read sequencing of plant genomes: An optimization of standard methods

IF 2.7 3区生物学 Q2 PLANT SCIENCES

Applications in Plant Sciences Pub Date : 2023-06-13 DOI:10.1002/aps3.11528

Myoungbo Kang, Andre Chanderbali, Seungyeon Lee, Douglas E. Soltis, Pamela S. Soltis, Sangtae Kim

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引用次数: 5

Abstract

Premise

Developing an effective and easy-to-use high-molecular-weight (HMW) DNA extraction method is essential for genomic research, especially in the era of third-generation sequencing. To efficiently use technologies capable of generating long-read sequences, it is important to maximize both the length and purity of the extracted DNA; however, this is frequently difficult to achieve with plant samples.

Methods and Results

We present a HMW DNA extraction method that combines (1) a nuclei extraction method followed by (2) a traditional cetyltrimethylammonium bromide (CTAB) DNA extraction method for plants with optimized extraction conditions that influence HMW DNA recovery. Our protocol produced DNA fragments (percentage of fragments >20 kbp) that were, on average, ca. five times longer than those obtained using a commercial kit, and contaminants were removed more effectively.

Conclusions

This effective HMW DNA extraction protocol can be used as a standard protocol for a diverse array of taxa, which will enhance plant genomic research.

Abstract Image

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用于植物基因组长读测序的高分子量DNA提取:标准方法的优化

开发一种有效且易于使用的高分子量(HMW) DNA提取方法对于基因组研究至关重要，特别是在第三代测序时代。为了有效地利用能够产生长读序列的技术，最大限度地提高提取DNA的长度和纯度是很重要的;然而，这通常很难在植物样本中实现。方法与结果提出了一种HMW DNA提取方法，该方法结合了(1)核提取法和(2)传统的十六烷基三甲基溴化铵(CTAB)提取植物DNA的方法，并优化了影响HMW DNA回收率的提取条件。我们的方案产生的DNA片段(片段百分比> 20kbp)平均比使用商业试剂盒获得的DNA片段长约5倍，并且更有效地去除污染物。结论该提取方案可作为植物基因组研究的标准方案，为进一步开展植物基因组研究奠定基础。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Applications in Plant Sciences PLANT SCIENCES-

CiteScore

7.30

自引率

0.00%

发文量

审稿时长

12 weeks

期刊介绍： Applications in Plant Sciences (APPS) is a monthly, peer-reviewed, open access journal promoting the rapid dissemination of newly developed, innovative tools and protocols in all areas of the plant sciences, including genetics, structure, function, development, evolution, systematics, and ecology. Given the rapid progress today in technology and its application in the plant sciences, the goal of APPS is to foster communication within the plant science community to advance scientific research. APPS is a publication of the Botanical Society of America, originating in 2009 as the American Journal of Botany''s online-only section, AJB Primer Notes & Protocols in the Plant Sciences. APPS publishes the following types of articles: (1) Protocol Notes describe new methods and technological advancements; (2) Genomic Resources Articles characterize the development and demonstrate the usefulness of newly developed genomic resources, including transcriptomes; (3) Software Notes detail new software applications; (4) Application Articles illustrate the application of a new protocol, method, or software application within the context of a larger study; (5) Review Articles evaluate available techniques, methods, or protocols; (6) Primer Notes report novel genetic markers with evidence of wide applicability.