中国兰花苞基因组DNA的提取及优化

Extraction and optimization of genomic DNA from Chinese orchid flower bud

采用改良CTAB法提取国兰花苞基因组DNA。优化部分如下:(1)在裂解细胞核前加入洗净液(0.4 mol·L^(-1)葡萄糖+3%可溶性PVP+10 mmol·L^(-1)β-巯基乙醇);(2)抽提时加入1/4体积的无水乙醇;(3)用等体积异丙醇沉淀DNA的同时加入1/2体积NaCl(5 mol·L^(-1))。结果表明,采用该法提取的DNA浓度介于216.5～245.8 ng·μL^(-1)之间,1.78<D_(260 nm)/D_(280 nm)<1.81,浓度和纯度均能满足常规分子生物学研究的需求。

The genomic DNA of Chinese orchid flower bud was extracted by an optimized CTAB method. The optimization included the following:(1) add a washing solution (0.4 mol·L^-1 glucose, 3% soluble PVP, 10 mmol·L^-1 β-mercaptoethanol) before lysing the nuclei;(2) add 1/4 volume of absolute ethanol;(3) while precipitating DNA with an equal volume of isopropanol, add 1/2 volume of 5 mol·L^-1 NaCl simultaneously. The results showed that the DNA concentration extracted by this method was greater between 216.5 and 245.8 ng·μL^-1 , the D 260 nm / D 280 nm ratio was between 1.78 and 1.81, both the purity and the concentration of genomic DNA met with the requirements of normal molecular biology experiments.