| 摘要: |
| 以裙带菜(Undaria pinnatifida)配子体为材料对比分析了4 种方法, 改进SDS 法、CTAB 法、Trizol试剂盒、RNAiso Reagent 试剂盒法提取RNA的质量和纯度, 并采用RT-PCR 对mRNA反转录, 对mRNA的可用性进行了检测。结果表明, 两种试剂盒Trizol 和RNAiso Reagent 及其改进的方法无法得到完整的裙带菜配子体总RNA; CTAB 法提取的总RNA 条带清晰完整但有明显的基因组DNA 残留并有多糖和蛋白质污染; 改进SDS 法提取的总RNA 三条带清晰, OD 值介于1.8～2.0 之间, 可应用于后续的RT-PCR 实验, 是一种较为理想的裙带菜等褐藻类配子体RNA 提取的方法。该方法用于其他大型海藻的总RNA 提取, 效果也比较好。 |
| 关键词: 裙带菜(Undaria pinnatifida) 配子体 RNA 提取 RT-PCR |
| DOI： |
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| 基金项目:中国科学院实验海洋生物学重点实验室开放基金项目 |
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| Comparison of methods for total RNA extraction from Undaria pinnatifida gametophyte |
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| Abstract: |
| In this paper, modified SDS method, CTAB method, and Trizol and RNAiso Reagent kits were applied to extract total RNA from Undaria pinnatifida gametophyte. We compared the quality and purity of the extracted RNAs and used RT-PCR to examine whether the RNAs were functionally active. The results showed that the Trizol and RNAiso Reagent kits were not suitable to seaweed RNA isolation, because there was no clear band detected on the gel. The RNA isolated by the CTAB method showed smear bands and also was contaminated by genomic DNA. However, the modified SDS method could give birth to good quality RNAs, although there was a little genomic DNA contamination. The OD260/OD280 ratio of the RNA was between 1.8 and 2.0. These RNAs could be used for RT-PCR. So the modified SDS method is a very good method for extracting total RNAs from gametophyte of U. pinnatifida and some other seaweeds. |
| Key words: Undaria pinnatifida gametophyte RNA extraction RT-PCR |
