香蕉不同组织的基因组DNA制备方法的比较

Comparison of Genomic DNA Extraction Methods from Various Tissues of Banana

本研究以香蕉的叶、假茎、根、果皮、果肉为材料,比较了CTAB常规法及其3种改良法、SDS法、SDS结合蛋白酶裂解法、改良尿素法、氯化苄法等方法提取基因组DNA的效果,以期筛选出通用性高、操作简便的基因组DNA制备方法。结果表明:氯化苄法不仅能够有效地从叶片、假茎、根系和果肉中提取到高质量的基因组DNA,也适合从果皮中提取少量DNA,其中提取液中添加600μL氯化苄的浓度处理效果最好;尽管CTAB常规法及其改良法能有效地从叶片中提取DNA,但对其他组织部位的提取效果较氯化苄法差;SDS法、SDS结合蛋白酶裂解法和改良尿素法的提取叶片DNA的效果较CTAB法差,也不能从根系和果皮、果肉中提取到高质量DNA。此外,PCR扩增试验结果表明,氯化苄法能满足后续的基于ITS的分子标记鉴定研究。

Abstrac t In order to select a simple and rapid protocol for high-throughput genomic DNA extraction from various tissues of banana（leaf, pseudostem, root, peel and pulp）, the eight existing methods, i.e, traditional or modified CTAB methods, SDS or SDS-proteinase K method, modified Urea method and Benzyl Chloride method,have been used to compare quality and efficiency respectively. Results showed that, Benzyl Chloride method not only facilitated the extraction of high quality DNA from tissues of leaf, pseudostem, root, pulp, but also was suitable to extract a small amount of DNA from peel after using an addition of 600 μL benzyl chloride in extraction solution. For traditional or modified CTAB methods, they exhibited less effective than Benzyl Chloride method,although the CTAB methods also performed well in extracting leaf DNA. Besides, genomic DNA could not been extracted from tissues of root, peel and pulp if using the methods of SDS, SDS-proteinase K or modified Urea, and the DNA quality from leaf extracted by these 3 methods was worse than those extracted by CTAB methods. In addition, the DNA extracted by Benzyl Chloride method was proved to be suitable for the downstream application,such as in PCR amplification of ITS region used for identifying and classifying banana germplasm.