海洋微藻活体及乙醇固定状态下基因组DNA的微量提取方法

探讨了海洋微藻活体及固定状态下基因组DNA的提取方法,即作者改进的CTAB法.结果表明,CTAB改进法提取的基因组DNA蛋白质、酚、盐和小分子的污染较少,多糖成份也得到了有效去除.用5%乙醇固定样本提取DNA的效果最好,对原生动物抑制效果也较明显;10%以上的乙醇可杀死原生动物,但对DNA的降解率较大;30%以上的乙醇对细菌抑制较明显,但DNA都有较大程度的降解.本法提取的DNA可用于正常的酶切和PCR.用甲醛、甲醇/冰醋酸等其它固定液固定样品,DNA的得率低或者质量和纯度低.图7表1参27

Genome DNA Extraction from Living and Alcohol-fixed Marine Micro-algae

Based on the method of CTAB with some modifications, genome DNA extraction of marine micro-algae from living and fixed samples was studied. The results indicated that the pollutions of genome DNA caused by protein, hydroxybenzene, salt and small molecules were lower, and amylose was eliminated efficiently by this extraction. The quality and quantity of DNA extracted from samples fixed in 5 alcohol was the best, and the activity of protozoan was restrained effectively. Protozoan in the samples fixed in 10 alcohol could be killed, but the quantity of the obtained genome DNA was lower. Reproduction of bacteria could be restrained effectively in the samples fixed in alcohol, of which the concentration exceeded 30 but the quantity of genome DNA was greatly decreased. The extracted DNA could be used in restriction endonuclease digestion and polymerase chain reaction. The yields of DNA extracted from the samples fixed in alcohol with other concentrations decreased respectively. Fixation with formalin and methanol acetic acid (MAA) resulted in poor yields or low quantity and purity of the extracted DNA. Fig 7, Tab 1, Ref 27