[关键词]
[摘要]
目的 采用SCoT分子标记方法对血叶兰Ludisia discolor及其近缘属植物资源进行亲缘关系及DNA指纹图谱构建,从分子水平对血叶兰及其近缘属种质进行区分。方法 使用正交设计方法,对SCoT-PCR反应体系进行优化,筛选适合血叶兰的反应体系、最佳退火温度及SCoT引物。以供试的12份种质资源为材料,利用筛选的引物对其进行分析,用POPgene计算材料间的遗传多样性系数,NTSYS软件进行聚类分析,并构建DNA指纹图谱。结果 获得适用于血叶兰的SCoT-PCR反应体系,筛选出12条多态性好的血叶兰SCoT引物,多态性比率为98.98%。根据Nei’s遗传相似系数,在相似系数为0.45时,12份种质资源可被分为3个分支。其中,斑叶兰与7份血叶兰种质归为I类,3份金线兰种质为Ⅱ类,而绿石蚕单独聚为Ⅲ类。利用SC8引物构建的DNA指纹图谱即可将12份材料鉴别开。结论 12份血叶兰及其近缘属种质具有丰富的遗传多样性,构建的DNA指纹图谱可用于血叶兰及其近缘属品系鉴定。
[Key word]
[Abstract]
Objective To make a distinction between Ludisia discolor and its relatives genus in molecular level, SCoT markers were employed to assess the genetic relationship and construct the DNA fingerprint. Methods Orthogonal design method were carried out to optimize the suitable SCoT-PCR reaction system based on five factors. The optimum annealing temperature and SCoT primers were also screened. The 12 germplasm resources were used as materials, the screened primers were selected to analyze the genetic relationship of 12 materials. POPGENE was used to calculate the genetic diversity, NTSYS was performed to analyze cluster, and DNA map was constructed. Results The optimized SCoT-PCR reaction system was constructed and a total of 12 rich bands were screened out as the primers of SCoT molecular marker with polymorphism ratio of 98.98%. According to Nei's genetic similarity coefficient, a total of 12 materials were divided into three cluster when coefficient was 0.45. Goodyera schlechtendaliana was in category I with seven L. discolor lines, indicating that these samples had close relationship. In category Ⅱ, there were three samples came from Anoectochilus roxburghii. Moreover, a green L. discolor sample was alone clustered into category Ⅲ. The DNA fingerprint map by the SC8 primer could identify the 12 materials. Conclusion There are rich genetic diversities in 12 samples of L. discolor and its relatives genus, and the construction of DNA fingerprint map provides a theoretical basis for the identification of L. discolor and its relatives genus, which were tested in this study.
[中图分类号]
R282.12
[基金项目]
漳州市科技重大专项项目(zz2018ZD20);广东省现代农业科技攻关项目(DzxQY008);福建省星火项目(2019S0031);福建省林业科技推广项目(2019TG08)