摘要
通过PCR引物检测人参DNA特异性片段,快速、准确的评价低龄人参种质质量,加快人参育种进程。该研究基于个体之间的基因差异,设计7对引物,通过比对不同人参DNA的扩增图谱与皂苷含量相关性,成功得到一对特异性引物GC1,并采用L_(16)(4~5)正交试验优化其25μL PCR体系,各组分最佳配比为:DNA 2.60 mg·L^(-1),Mg2+1.44 mmol·L^(-1),d NTP 0.19mmol·L^(-1),引物0.32μmol·L^(-1),Taq 0.076 U·μL^(-1)。对GC1引物进行验证,在24株供试人参中有2株扩增出1 200 bp特异性条带阳性样品,2株阳性样品中9种单体皂苷及其加和值含量均较高。该特异性条带序列与甘油-3-磷酸脱氢酶同源性达99%。GC1可以作为一种高皂苷人参株系早期筛选鉴定方法的特异性引物,有利于加快人参育种进程。
The study aims at screening the specific bands by PCR, quickly and accurately evaluating the quality of ginseng seeding, accelerating the process of ginseng breeding. Based on the correlation of genetic differences and saponin content between individuals, a pair of specific primer GC1 was screened by PCR. According to the experiment by L16 （45） orthogonal test, a PCR system most suitable for GC1 was established, which came out total 25 μL reaction system containing DNA 2.60 mg·L^-1, Mg2＋ 1.44 mmol·L^-1, dNTP 0.19 mmol·L^-1, primer 0.32 μmol·L^-1and Taq enzyme concentration 0.076 U·μL^-1. By comparing the saponin content and the GC1 PCR electrophoretogram of samples, the ginseng, with 1 200 bp specific band by PCR of GC1, the contents of 9 monosodium saponins and their additions were higher than others, which provided a reliable method for accelerating the process of ginseng breeding. The sequence was sequenced and 99% homologous to glycerol-3-phosphate dehydrogenase.
出处
《中国中药杂志》
CAS
CSCD
北大核心
2017年第24期4775-4781,共7页
China Journal of Chinese Materia Medica
基金
国家重点研发计划项目(2016YFC0500300)
吉林省重点科技成果转化项目(20160307005YY
20170204017YY)
关键词
人参育种
PCR
引物筛选
体系优化
ginseng breeding
PCR
primer screening
system optimization
