条斑紫菜高纯度总DNA及其质粒状DNA的提取
Extraction of plasmid-like DNA and high-quality total DNA from Porphyra yezoensis
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摘要: 提取条斑紫菜高纯度总DNA及其质粒状DNA的新方法。先用海螺酶处理紫菜叶状体制备细胞,然后用SDS-蛋白酶K裂解细胞提取总DNA,再用玻璃粉浆(glassmilk)对其纯化,经纯化后的总DNA能被EcoRI,Dral与HaeⅢ等限制酶完全酶切,并在酶切图谱上形成明显的DNA带型。当用异硫氰酸胍一十二烷基肌氨酸钠裂解紫菜细胞时,在总DNA提取物中直接发现有一条质粒状DNA带(2.3Kb),即建立了一种极简便的质粒状DNA提取方法。Abstract: Cellls were prepared from sea snail enzyme digests of Porphyra yezoensis Thalli. Crude extracts of total DNA were isolated from Porphyra cells lysed with SDS-Proteinase K solution. Total DNA was purified with glassmilk, the resulting DNA was of high quality, could be completely digested with restriction enzymes. So that it was also suitable for analyses of Southern hybridization, RFLP and AFLP, etc. DNA bands were clearly observed in the agarose gel electrophoresis patterns of total DNAs digested with EcoRI, PstI and HaeⅢ respectively. The presence of DNA bands in the electrophoresis patterns of total DNA showed that the nuclear genome of Porphyra yezoensis may be small. Unexpectedly, a plasmid-like DNA band (2.3Kb) was found in the alectrophoresis pattern of P. Yezoensis total DNA isolated with guanidinium thiocyanate-sarcosyl solution. This is a very simple and convenient method for plasmid-like DNA isolation.
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Key words:
- Porphyra yezoensis /
- cells /
- total DNA /
- glassmilk /
- complete digestion /
- plasmid-like DNA
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