Liu Huan, Zhao Haixia, Wu Longhua, Xu Wenzhong
Key Laboratory of Plant Resources, Institute of Botany, Chinese Academy of SciencesBeijing, China.
University of Chinese Academy of SciencesBeijing, China.
Front Plant Sci. 2017 Jun 16;8:1047. doi: 10.3389/fpls.2017.01047. eCollection 2017.
The present study demonstrates the development of an -mediated genetic transformation method for species of the genus, which includes the Cd/Zn hyperaccumulator and the non-hyperaccumulating ecotype of . Multiple shoots were induced from stem nodes of two plants using Murashige and Skoog (MS) medium containing 0.1 mg/L cytokinin 6-benzyladenine (6-BA) and 1.0 mg/L auxin 1-naphthaleneacetic acid (NAA). The shoot primordia were used as direct targets for infection. Selection on hygromycin was highly effective in generating -transformed explants. This callus-free procedure allowed us to obtain transgenic plantlets after rooting hygromycin-resistant shoots on phytohormone-free MS medium containing the antibiotic. The presence and expression of the reporter genes and in transgenic plants were confirmed by a real-time polymerase chain reaction, histochemical GUS assays, and confocal microscopy. This reliable method for genetic transformation of plants will help us to understand gene functions and the molecular mechanisms underlying Cd hypertolerance and hyperaccumulation in these species.
本研究展示了一种用于该属物种的农杆菌介导的遗传转化方法的开发，该属包括镉/锌超积累植物以及该植物的非超积累生态型。使用含有0.1 mg/L细胞分裂素6-苄基腺嘌呤（6-BA）和1.0 mg/L生长素1-萘乙酸（NAA）的Murashige和Skoog（MS）培养基，从两种植物的茎节诱导出多个芽。芽原基被用作感染的直接靶标。在潮霉素上进行选择对于产生农杆菌转化的外植体非常有效。这种无愈伤组织的方法使我们能够在含有抗生素的无植物激素的MS培养基上使抗潮霉素的芽生根后获得转基因小植株。通过实时聚合酶链反应、组织化学GUS分析和共聚焦显微镜证实了转基因植物中报告基因的存在和表达。这种可靠的植物遗传转化方法将有助于我们了解这些物种中基因功能以及镉耐受性和超积累的分子机制。
