| 摘要: |
| 用室内培养的塔玛亚历山大藻(Alexandrium tamarense ATHK藻株)作为实验对象,分别选择了5%福尔马林固定后常温保存、95%乙醇固定后常温保存、- 20℃冷冻保存、鲁哥氏液(Lugolps solution)固定后常温保存等方法,在5,15,30,60d后对保存的样品进行单细胞PCR反应,并与新鲜样品进行比较。实验结果表明,用95%乙醇保存、-20℃冷冻保存、鲁哥氏液保存的样品在60d后仍可扩增出目标条带,与新鲜样品没有差别,但是样品中的藻细胞形态有一些改变;而以5%福尔马林固定保存的样品只能在5d后扩增出目标条带,但藻细胞形态比较完好。因此,对用于单细胞PCR实验的微藻样品,短期内(不多于5d)可以采用福尔马林保存,而需要长期保存的样品,应当采用乙醇或鲁哥氏液固定,或者采用-20℃冷冻保存的方法。 |
| 关键词: 有害赤潮 单细胞PCR 保存方法 塔玛亚历山大藻(Alexandrium tamarense) |
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| 基金项目:国家自然科学基金项目( 4067072) ; 国家863 计划项目(2006AA09Z178) |
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| Comparison of sample preservation methods in single-cell PCR experiment of algae |
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| Abstract: |
| Recently, more and more molecular biological methods and techniques have been used in harmful-algal-bloom (HAB) studies. The single-cell PCR method, for example, is promising in genetic studies of HAB species which is hard to culture in the lab. However, little information is available on preservation of the algae samples used for single-cell PCR study. In this experiment, the cultured cells of Alexandrium tamarense were used to compare the different sample preservation methods, including formalin fixation (5%), ethanol fixation (95%), Lugolps solution fixation, and refrigeration at -20℃,to find the appropriate sample preservation method. The valid time for each sample preservation method was also tested, using samples preserved for 5d, 15d, 30d and 60d, respectively. It was found that algae samples preserved by ethanol fixation (95%), Lugol’s solution fixation or refrigeration at -20℃ for as long as 60d could still be effectively used for single-cell PCR study. However, for formalin fixation (5%) method, only sample preserved for 5d gave a positive single2cell PCR result. The morphological features of the algae samples were well preserved by formalin fixation (5%). These results suggest that formalin fixation (5%) is a good choice for short-term (less than 5d) preservation of algae samples, while 95% ethanol fixation, Lugolps solution fixation, or refrigeration at -20℃are suitable for relatively long-term preservation of algae samples. |
| Key words: harmful algal bloom single-cell PCR Preservation Alexandrium tamarense |
