收稿日期: 2009-12-10
修回日期: 2009-12-24
网络出版日期: 2010-12-15
基金资助
广西科学基金项目(桂科基0575008); 广西大学基金项目(X041120)
DNA isolation and PCR amplified in Gracilaria
Received date: 2009-12-10
Revised date: 2009-12-24
Online published: 2010-12-15
Supported by
广西科学基金项目(桂科基0575008); 广西大学基金项目(X041120)
通过对CTAB法、SDS法和植物基因组提取试剂盒3种方法提取江蓠Gracilaria总DNA的得率比较, 并将得到的总DNA用于限制性酶切和PCR扩增反应进行纯度检测, 选择出适合江蓠属总DNA的提取方法, 建立了江蓠RAPD、ISSR和ITS等遗传多样性和系统学研究的分子生物学方法。实验结果表明, SDS法和植物基因组提取试剂盒均适用于江蓠总DNA的提取; 其中SDS法适用于新鲜和冰冻材料的总DNA提取, 不仅得率和纯度高, 且方法稳定性好, 提取时间较短, 在本实验中是江蓠总DNA提取的最佳方法。植物基因组提取试剂盒的得率低, 但质量好, 操作快速简便, 适用于大量新鲜样本总DNA的提取。CTAB法由于相对耗时长且产物稳定性差, 不推荐在江蓠中使用。
关键词： 江蓠Gracilaria; 总DNA提取; PCR扩增
李文红,房振峰 . 江蓠总DNA提取及其PCR扩增方法的建立[J]. 热带海洋学报, 2010 , 29(6) : 98 -103 . DOI: 10.11978/j.issn.1009-5470.2010.06.098
The proper total DNA isolation methods of Gracilaria were chosen from three popular methods known as CTAB, SDS, and commercial Plant DNA Extraction kits by comparing their yield, quality, reliability, and the extraction time con-sumed. The total DNA was applied to restricted enzyme digestion and PCR amplification to detect its quality. The reactions of ingredients and programs of RAPD, ISSR, and ITS were constructed based on the detections. The results showed that the SDS method is the best method among the three for its highest yield and fine quality of total DNA, both good for fresh and frozen Gracilaria samples, not time-consuming and excellent reliability. The Plant DNA Extraction Kit was quick and user-friendly for the mass fresh samples DNA isolation in Gracilaria though its total DNA yield was low. The CTAB method was not rec-ommended for its time-consuming and poor reliability.
Key words： Gracilaria; total DNA isolation; PCR
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