紫茎泽兰DNA的提取及AFLP反应体系的建立

Genomic DNA Extraction Method and AFLP Amplification Reaction System of Ageratina adenophora Spreng.

摘要: 以紫茎泽兰叶片为材料,用改进的CTAB法,在提取液中加入2%PVP40(V/V)、0.4%BME(V/V),提取到了高质量的基因组DNA,OD₂₆₀/OD₂₈₀在1.7~1.9之间,蛋白质、多酚类、多糖、RNA等去除较彻底,适于AFLP分析。通过对紫茎泽兰DNA提取、酶切连接、预扩增、选择性扩增等实验过程中各关键因素的比较研究,建立了一套优化的AFLP分子标记体系,得到了清晰的AFLP银染指纹图谱,实验结果重复性好。

Abstract: The study dealt with extraction of DNA with improved CTAB(cetyl-trimethyl-ammonium bromide) method from Ageratina adenophora Spreng.It showed that CTAB buffer supplemented with 2% PVP40(polyvinylpyrrolidone 40) and 0.4% β-mercaptoethanal was suitable for the genomic DNA extraction of A.adenophora,with OD₂₆₀/OD₂₈₀ value of 1.7-1.9,representing high purity of the extracted DNA and the complete elimination of protein,phenol,polyhexose and RNA.Thus the improved CTAB method was approved suitable for further AFLP analysis of A.adenophora.Some key factors affecting DNA extraction,restriction-ligase reaction,pre-amplification and amplification processes were studied and an optimized AFLP reaction system of A.adenophora was established.With this AFLP reaction system,clear DNA electrophoresis fingerprints of A.adenophora was obtained,and the reproducibility was high.