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以6份不同生态区野生大麻为试验材料,对叶绿体17个高变区的通用引物进行多态性筛选,同时基于已公布的大麻叶绿体基因组全序列,应用生物信息学软件对其进行比较基因组学分析,开发新的多态性引物。结果显示:(1)17对通用引物,通用性达76.5%,但仅ndh F-rpl32引物产物多态性好;(2)比较基因组学研究表明大麻叶绿体基因组多态性比较低,但存在极少的高变区,针对全序列高变区段设计的4对引物(trn L-rpl32~a,rps8-rps11,psa I-acc D,rps16~a)在6份野生大麻群体材料中分别有2~8个变异位点,具有较高的多态性。叶绿体非编码基因片段通用引物ndh F-rpl32及本研究针对大麻开发的trn L-rpl32~a、rps8-rps11、psa I-acc D、rps16~a引物在大麻中具有专一性强、产物多态性高等特点,可作为高效分子标记应用于大麻种质资源遗传多样性与保护、物种起源与遗传进化、种下分类鉴定、工业大麻育种等研究。
Six wild cannabis samples from different ecotypes were used to screen 17 universal primers of chloroplast. Based on the published sequence of the genome of cannabis chloroplast genome, comparative genomics Analyze and develop new polymorphic primers. The results showed that: (1) 17 pairs of universal primers had a commonality of 76.5%, but only the polymorphism of ndh F-rpl32 primer was good; (2) Comparative genomics showed that the polymorphism of chloroplast genome was low, Four pairs of primers (trn L-rpl32-a, rps8-rps11, psa I-acc D, rps16-a) designed for the entire sequence of hypervariable regions were used in 6 wild populations of cannabis 2 to 8 mutation sites, with high polymorphism. Chloroplast non-coding gene fragment universal primer ndh F-rpl32 and development of this study for the development of cannabis trn L-rpl32 ~ a, rps8-rps11, psa I-acc D, rps16 ~ a primer in marijuana has strong specificity, It can be used as efficient molecular marker in genetic diversity and conservation of hemp germplasm resources, species origin and genetic evolution, species classification and identification, and industrial cannabis breeding.