High-molecular-weight and pure DNA is crucial for high-quality results from 3rd generation DNA Analyzers and optical mapping technologies. Conventional nuclei isolation methods for preparing high-molecular-weight genomic DNA from plant tissues include the preparation of protoplasts or embedding nuclei in an agarose matrix with subsequent manipulations via electro-elution or pulsed-field gel electrophoresis. In this method, plant nuclei are isolated by physically grinding tissues and reconstituting the intact nuclei in a unique Nuclear Isolation Buffer (NIB). The plastid DNAs are released from organelles and eliminated with an osmotic buffer by washing and centrifugation. The purified nuclei are then lysed and further cleaned by organic extraction, and the genomic DNA is precipitated with a high concentration of CTAB. The highly pure, high molecular weight gDNA is extracted from the nuclei, dissolved in a high pH buffer, allowing for stable long-term storage. This method is unique and avoids the use of embedding in agarose, which dramatically reduces time (4–8 h versus days), complexity, and materials cost. This procedure can be used on essentially any plant species and tissue stage. Here we describe a case study and a simple method to rapidly prepare high molecular weight gDNA from Upland cotton, blackgrass, and strawberry suitable for single-molecule sequencing.

中文翻译：

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适用于单分子技术的简单植物高分子量 DNA 提取方法

高分子量和纯 DNA 对于第三代 DNA 分析仪和光学作图技术的高质量结果至关重要。从植物组织中制备高分子量基因组 DNA 的常规细胞核分离方法包括制备原生质体或将细胞核嵌入琼脂糖基质中，随后通过电洗脱或脉冲场凝胶电泳进行操作。在这种方法中，通过物理研磨组织并在独特的核分离缓冲液 (NIB) 中重组完整的细胞核来分离植物细胞核。质体 DNA 从细胞器中释放出来，并通过洗涤和离心用渗透缓冲液消除。然后将纯化的细胞核裂解并通过有机萃取进一步清洁，并用高浓度的 CTAB 沉淀基因组 DNA。高纯度，从细胞核中提取高分子量 gDNA，溶解在高 pH 值缓冲液中，从而实现稳定的长期储存。这种方法是独一无二的，避免了在琼脂糖中嵌入的使用，从而大大减少了时间（4-8 小时与天）、复杂性和材料成本。该程序可用于基本上任何植物物种和组织阶段。在这里，我们描述了一个案例研究和一种从陆地棉、黑草和草莓中快速制备适合单分子测序的高分子量 gDNA 的简单方法。该程序可用于基本上任何植物物种和组织阶段。在这里，我们描述了一个案例研究和一种从陆地棉、黑草和草莓中快速制备适合单分子测序的高分子量 gDNA 的简单方法。该程序可用于基本上任何植物物种和组织阶段。在这里，我们描述了一个案例研究和一种从陆地棉、黑草和草莓中快速制备适合单分子测序的高分子量 gDNA 的简单方法。