一种从果树成熟叶片提取DNA的方法

针对果树成熟叶片富含多糖、多酚及其它次生代谢物质的特点,以8个果树树种的秋季成熟叶片为材料,采用改进的CTAB法对其基因组总DNA进行了提取,并对得到的DNA进行了电泳检测、含量测定和AFLP分析。结果表明,该方法从8种果树的成熟叶片中均能提取出基因组总DNA,且DNA的纯度和完整性都较好,D260nm/D280nm的比值均在1.8～2.0之间,无降解现象,RNA去除干净,能被限制性内切酶完全消化;其AFLP分析(引物组合为EcoRI-TA/MseI-CTT,EcoRI-TT/MseI-CTT)条带清晰,多态性好,说明此方法提取的果树成熟叶片基因组总DNA完全适于AFLP分析。

A method for DNA extraction from mature leaves of fruit trees

The mature leaves of fruit tree are rich in polyhexose, polyphenols and other secondary metabolism compounds which prevent DNA extraction. The genomic DNA samples of mature leaves of 8 fruit trees in autumn were isolated by improved CTAB method, and the purity and quantity of each DNA obtained was evaluated by agarose gel electrophoresis, UV scanning and AFLP analysis. The results showed, each of the genomic DNA extracted from mature leaves of 8 fruit trees by this method was pure, integral, the value of D260 nm/D280 nm was 1.80 to 2.0, degradation was nearly free, the RNA was eliminated completely, and suitable for digestion by restriction endonucleases. AFLP analysis indicated the primer (EcoRI-TA/MseI-CTT, EcoRI-TT/MseI-CTT) produced clear polymorphic patterns. Therefore this method could be used for extracting ideal DNA samples from mature leaves of fruit trees and suitable for AFLP.