厚朴AFLP分子标记体系的建立

An AFLP molecular labeling technique for Magnolia officinalis

以厚朴Magnolia officinalis为材料,通过对扩增片段长度多态性(AFLP)反应体系中的酶切模板用量、扩增条件等几个方面进行优化,建立了较为理想的厚朴AFLP反应体系。结果表明:①采用十六烷基三甲基溴化铵(CTAB)硅珠法提取的DNA质量较好,适用于AFLP分析。②酶切的基因组DNA以400ng为宜;预扩增产物的稀释倍数以30倍为宜;20.00μL选扩反应体系中Mg2+最佳浓度为2.00mmol·L-1,三磷酸脱氧核苷酸(dNTPs)最佳浓度为0.225mmol·L-1,Taq DNA聚合酶最佳用量为16.67nkat时,扩增带清晰且稳定,扩增效果较好。旨在为研究厚朴天然种群的遗传多样性及遗传结构分析、新品种选育及鉴定和遗传图谱构建等奠定理论基础。

To establish an efficient and stable amplified fragment length polymorphism （AFLP） analysis system for Magnolia officinalis, selection and optimization of DNA template treatments and selective amplification conditions were undertaken using the cetyl trimethylammonium bromide （CTAB）-SiO2 method for mapping. Results indicated that： High quality DNA samples were obtained with the CTAB-SiO2 method and the DNA template used was 400 ng. The best treatments for the analysis were Mg2＋ of 2.00 mmol·L-1, de oxyribonucleotide triphosphates （dNTPs） of 0.90 mmol·L-1, and Taq DNA polymerase of 16.67 nkat. This research showed that the CTAB-SiO2 method was fit for AFLP analysis on Magnolia, that a 30 times dilution for the pre-amplification products could be used for selective amplification, and that the silver-staining AFLP system could provide a method for further study with molecular markers in the field of genetic diversi- ty, for breeding new cuhivars, and for genetic relationships with Magnolia officinalis. [Ch, 7 fig. 1 tab. 12 ref. ]