相关实验视频

Updated: May 23, 2025

Optimization and Comparative Analysis of Plant Organellar DNA Enrichment Methods Suitable for Next-generation Sequencing

Optimization and Comparative Analysis of Plant Organellar DNA Enrichment Methods Suitable for Next-generation Sequencing

Published on: July 28, 2017

改进的提取方法将高分子量DNA从Magnaporthaceae和其他草根真菌中分离出来,用于长读全基因组测序

Michelle J Grey¹, Jackie Freeman², Jason Rudd²

¹Department of Organisms and Ecosystems, Earlham Institute, Norwich, England.

|May 21, 2025

概括

经过修改的协议有效地将高分子量 (HMW) DNA从Magnaporthaceae真菌中分离出来,用于全基因组测序. 这些方法提高了DNA完整性和产量,这对于准确的长读测序结果至关重要.

科学领域:

分子生物学分子生物学
菌类学菌类学是指菌类学.
基因组学就是基因组学.

背景情况:

高分子量 (HMW) DNA对于精确的全基因组测序至关重要,特别是对于复杂的基因组.
的二次代谢物,如黑色素,可以干扰DNA提取和测序.
现有的DNA提取协议可能不适合所有真菌物种,特别是那些有色素问题的人.

研究的目的:

开发和优化修改后的协议,以从Magnaporthaceae真菌菌中分离HMWDNA.
适应现有的商业DNA提取套件 (Cytiva Nucleon PhytoPure和Macherey-Nagel NucleoBond HMW),以提高HMW DNA的产量和完整性.
确保可靠的HMW DNA适合长读全基因组测序.

主要方法:

修改了Cytiva Nucleon PhytoPure套件:增加了RNase A处理,添加了蛋白酶K,四倍提取.
适应Macherey-Nagel NucleoBond HMW套件:样品研磨,一夜间溶解,洗洗前的DNA卷轴.
基于生长速度和二次代谢物积累的样本分类.

主要成果:

修改后的Cytiva套件从测试的一半物种中恢复了HMW DNA.
经过修改的马切里-纳格尔套件使用较温和的重力柱产生了>50%的高纯度DNA链>40kbp.
两种协议都优先考虑在洗前将DNA卷起来的DNA完整性.

结论:

开发的协议提供了可靠的方法,从Magnaporthaceae真菌中获得高质量的HMWDNA.
这些优化的方法适用于长时间读取的全基因组测序,增强下游分析.
该方案可适应其他ascomycetes,可在四个样本 (不包括溶解) 中大约4小时内完成.

关键词:

菌的真菌是一种在HMW DNA提取过程中,长读序列的测序方式马格纳普尔太家族 (Magnaporthaceae) 的一个家族.整个基因组的整个基因组.

更多相关视频

Purification of High Molecular Weight Genomic DNA from Powdery Mildew for Long-Read Sequencing

Purification of High Molecular Weight Genomic DNA from Powdery Mildew for Long-Read Sequencing

Published on: March 31, 2017

Extraction of High Molecular Weight Genomic DNA from Soils and Sediments

Extraction of High Molecular Weight Genomic DNA from Soils and Sediments

Published on: November 10, 2009

相关概念视频

DNA Isolation

DNA Isolation

DNA isolation protocols can be fast and straightforward or complex and time-consuming depending on the type and quality of DNA required for further processing. For example, plasmid DNA extraction is a bit more complicated than genomic DNA extraction because of the need for an appropriate lysis method to separate plasmid DNA from gDNA during isolation. However, for specific applications, such as long-range DNA sequencing that require a good yield of high- quality DNA samples, we need to follow...