Preparation and purification of DNA from insects for AFLP analysis
Preparation and purification of DNA from insects for AFLP analysis
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DOI：
10.1046/j.1365-2583.1998.71048.x
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发表时间：
1998-02-01
影响因子：
2.6
通讯作者：
Zebitz, CPW
中科院分区：
文献类型：
--
作者：
Reineke, A;Karlovsky, P;Zebitz, CPW
扩增片段长度多态性（AFLP）分析有可能成为研究节肢动物遗传关系和遗传多样性的一种强有力的新DNA指纹技术。由于高质量的DNA是AFLP分析的一个重要先决条件，我们评估了六个协议（一个快速和四个复杂的方法与酚-氯仿处理，以及一个CTAB为基础的方法）的适用性提取DNA从昆虫材料和三个额外的DNA纯化步骤。最快速的DNA提取方法不能产生适合AFLP分析的DNA。在测试的四种复杂方法中，两种方案导致DNA产量相对较低，因此不能用作AFLP分析的模板。另外两个复杂的方法与苯酚处理和CTAB为基础的DNA提取协议提供DNA适合AFLP分析。使用精胺沉淀的DNA的额外纯化揭示了在AFLP凝胶中被未纯化的DNA掩盖的一些额外条带。因此，AFLP模板推荐使用精胺沉淀法。
Analysis of amplified fragment length polymorphism (AFLP) has the potential to become a powerful new DNA fingerprinting technique for studying genetic relationships and genetic diversity in arthropods. Since DNA of high quality is a crucial prerequisite for AFLP analysis we evaluated the applicability of six protocols (one fast and four complex methods with phenol-chloroform treatments as well as one CTAB-based method) for extracting DNA from insect material and three additional DNA purification steps. The most rapid DNA isolation method did not produce DNA suitable for AFLP analysis. Among four complex methods tested, two protocols resulted in comparatively low yields of DNA that was therefore not used as template for AFLP analysis. The other two complex methods with phenol treatments and a CTAB-based DNA extraction protocol provided DNA suitable for AFLP assay. An additional purification of the DNA using spermine precipitation revealed a few extra bands in an AFLP gel that were masked in unpurified DNA. Therefore spermine precipitation is recommended for AFLP templates.