Molecular Taxonomy Lab, Botany Division, Forest Research Institute (FRI), P.O. New Forest, Dehradun, Uttarakhand, India.
Biotechnol Lett. 2013 Jun;35(6):961-8. doi: 10.1007/s10529-013-1152-3. Epub 2013 Feb 10.
Among five hairy root lines of Picrorhiza kurrooa that were established through Agrobacterium rhizogenes, one (H7) was selected for encapsulation due to high accumulation of picrotin and picrotoxinin (8.3 and 47.6 μg/g DW, respectively). Re-grown encapsulated roots induced adventitious shoots with 73 % frequency on MS medium supplemented with 0.1 μM 6-benzylaminopurine, following 6 months of storage at 25 °C. Regenerated plantlets had 85 % survival after 2 months. Regenerants were of similar morphotype having increased leaf number and branched root system as compared to non-transformed plants. The transformed nature of the plants was confirmed through PCR and Southern blot analysis. Genetic fidelity analysis of transformed plants using RAPD and ISSR showed 5.2 and 3.6 % polymorphism, respectively. Phytochemical analysis also showed that picrotin and picrotoxinin content were similar in hairy root line and its regenerants.
在通过发根农杆菌建立的 5 株罗锅底发根系中,由于 picrotin 和 picrotoxinin 的高积累(分别为 8.3 和 47.6μg/g DW),选择了 1 株(H7)进行包封。在添加 0.1μM 6-苄基氨基嘌呤的 MS 培养基上,重新生长的包封根在 25℃下储存 6 个月后,诱导不定芽的频率为 73%。经过 2 个月的培养,再生植株的存活率为 85%。与非转化植株相比,再生植株的形态相似,叶片数量增加,根系分枝。通过 PCR 和 Southern blot 分析证实了植株的转化特性。使用 RAPD 和 ISSR 对转化植株进行遗传保真度分析,分别显示出 5.2%和 3.6%的多态性。植物化学分析还表明,毛状根系及其再生体中的 picrotin 和 picrotoxinin 含量相似。