板栗基因组AFLP银染反应体系的建立

为了建立准确、高效、实用的板栗品种鉴别体系和方法,采用改进的CTAB—DNA提取方法,从板栗的嫩叶和老叶中提取获得总DNA。所得DNA样品的D260nm/D280nm值为1.7～1.9,琼脂糖凝胶上主带清晰、较少降解、样品纯度高、蛋白去除干净且获得率高。该体系使用操作简便、快捷、高效,适合于板栗DNA提取,所提取的DNA样品不含PCR反应抑制剂及其他酶反应抑制剂,可被限制性内切酶MseI和EcoRI完全酶切,适合AFLP-PCR反应应用,得到清晰的指纹式样,适宜用作板栗品种鉴别的有效方法。

Establishment of AFLP silver-stained reaction system for genomic DNA in Castanea mollissima

The improved method of CTAB-DNA isolation was used to extract genomic DNA from the tender or old leaves of Castanea mollissima. High quality total DNA was isolated in less than 3 hours. The values of D260 nm/D280 nm of each sample were between 1.7 and 1.9. Strips in agarose electrophoresis were clear and less decomposed. DNA samples with high purity and high yield could be gained by this method. The procedure of the method is easy,fast and effective,and is good to extract genomic DNA of Castanea mollissima. These genomic DNA samples were intact without contaminating nucleases or inhibitor. The DNA isolated with this method could be completely digested by restriction enzyme of Mse I and EcoR I, and good enough for use in following applications, including PCR and analysis of AFLP. The systems suitable for silver stained AFLP analysis were established and capable of producing sharp finger print patterns.