檀香心材总RNA提取方法的比较研究

Comparison of Several RNA Extracting Methods from Heartwood of Santalum album

摘要:

为筛选檀香心材总RNA提取方法，对5种提取方法进行比较研究，包括Trizol法、改良CTAB法、SDS酸酚法、异硫氰酸胍-CTAB法、异硫氰酸胍-SDS法。结果表明，Trizol法和异硫氰酸胍-CTAB法不能提取出檀香心材总RNA，而SDS酸酚法、改良CTAB法和异硫氰酸胍-SDS法均能提取檀香心材总RNA。SDS酸酚法的A₂₆₀ nm/A₂₃₀ nm小于2.0，且RNA产率低，仅为（27.94±1.06）μg g^-1，不能满足后续实验要求。而改良CTAB法和异硫氰酸胍-SDS法提取的总RNA带型清晰，完整性好，A₂₆₀ nm/A₂₈₀ nm为1.8~2.0，A₂₆₀ nm/A₂₃₀ nm大于2.0，RNA产率分别为（79.06±4.22）和（107.00±1.36）μg g^-1。分别以改良CTAB法和异硫氰酸胍-SDS法提取的总RNA为模板，通过RT-PCR反应，扩增檀香Actin基因片段，结果二者扩增产物大小相同且条带单一，说明改良CTAB法与异硫氰酸胍-SDS法为檀香心材总RNA提取的较好方法。

Abstract:

In order to screen RNA extracting method from heartwood of Santalum album, five methods were compared, including Trizol method, improved CTAB method, SDS-phenol method, guanidine isothiocyanate-CTAB method, and guanidine isothiocyanate-SDS method. The results showed that it was difficult to extract total RNA by Trizol method and guanidine isothiocyanate-CTAB method, and the improved CTAB method, SDS-phenol method and guanidine isothiocyanate-SDS method could successfully extract total RNA. The SDS-phenol method was not suitable for RNA extraction, because A₂₆₀ nm/A₂₃₀ nm was less than 2.0 and total RNA yield was low with (27.94±1.06) μg g^-1. However, the 28S rRNA and 18S rRNA bands of total RNA extracted by improved CTAB method and guanidine isothiocyanate-SDS method were clear, A₂₆₀ nm/A₂₈₀ nm ranged from 1.8 to 2.0, A₂₆₀ nm/A₂₃₀ nm were more than 2.0, the yields were (79.06±4.22) and (107.00±1.36) μg g^-1, respectively. The Actin segments in total RNA extracted by improved CTAB method and guanidine isothiocyanate-SDS method could amplificated with the same size and sigle band. Therefore, the improved CTAB methodand and guanidine isothiocyanate-SDS method were effective methods to extract total RNA from heartwood of S. album.