Method for increasing positive rate of transgenic Salvia miltiorrhiza

提高转基因丹参中阳性率的方法

  • Inventors: ZHU YUPAN
  • Assignees: 朱育盼
  • Publication Date: March 15, 2017
  • Publication Number: CN-106497969-A

Abstract

The invention relates to a method for increasing positive rate of transgenic Salvia miltiorrhiza. By employing Agrobacterium tumefaciens mediated leaf dish transformation, the method is simple to perform, high in repeatability, and effective in producing plants of transgenic Salvia miltiorrhiza quickly; at present, most Salvia miltiorrhiza plant genetic transformation systems are screened by using kanamycin; however, genetic transformation researches by the present lab on 99-3 variety of Salvia miltiorrhiza already subjected to sequencing discover that 99-3 Salvia miltiorrhiza leaf is sensitive to kanamycin, the growth of the leaf easily stops when kanamycin is used for screening antibiotics, and experimental reproducibility is poor. By screening with 50 mg/L Hyg, positive transgenic plants are acquired, with very high positive rate nearly 100%. The method is an efficient, reliable and convenient method for increasing positive rate of Salvia miltiorrhiza plants, where carrier foreign plasmids are transferred to Salvia miltiorrhiza explants through Agrobacterium tumefaciens genetic transformation, by using Salvia miltiorrhiza leaves as the explants.
本发明涉及一种提高转基因丹参中阳性率的方法,应用根癌农杆菌介导的叶盘转化法操作简便、重复性高、能快速、有效获得转基因丹参植株。目前丹参植株遗传转化体系多采用卡那霉素进行筛选,但本实验室在对已经完成测序工作的99‑3品系的丹参进行遗传转化研究中发现99‑3丹参叶片对卡那霉素敏感,用它做筛选抗生素时叶片生长容易停止,实验重现性差。本方法采用50mg/L Hyg进行筛选时,不仅获得了阳性转基因植株,而且阳性率极高,接近100%。上述方法建立了一种以丹参叶片作为外植体,通过根癌农杆菌的遗传转化法将携带外源质粒转入丹参外植体中的高效可靠、简便的丹参植株提高阳性率的方法。

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