An Efficient Procedure for Regeneration from Leaf-derived Calluses of Lonicera macranthoides 'Jincuilei', an Important Medicinal Plant
An Efficient Procedure for Regeneration from Leaf-derived Calluses of Lonicera macranthoides 'Jincuilei', an Important Medicinal Plant
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DOI：
10.21273/hortsci.44.3.746
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发表时间：
2009-06
期刊：
影响因子：
1.9
通讯作者：
Xiaoming Wang;Jianjun Chen;L. Yongxin;Q. Nie;Jun-bao Li
中科院分区：
文献类型：
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作者：
Xiaoming Wang;Jianjun Chen;L. Yongxin;Q. Nie;Jun-bao Li
“金翠蕾”是从灰毡毛忍冬中筛选出的一个突变体。马兹它产生丰富的花朵，永远不会开放，绿原酸（CGA）含量高达6.0%。通过生根或嫁接繁殖只有30%的成活率。本研究旨在建立一种快速再生该突变体的有效方法。叶片外植体接种在Gamborg's B 5培养基上，附加不同浓度的6-苄基腺嘌呤（BA）和2，4-二氯苯氧乙酸（2，4-D）。最佳的诱导组合为4.4 μM BA +2.26 μ m2，4-D，在4周内叶片外植体产生愈伤组织的比例为86.7%。将诱导出的愈伤组织接种在含有不同浓度激动素（KT）和α-萘乙酸（NAA）的B 5培养基上。诱导芽的最佳配方为B 5 + 0.9 μM KT +5.4 μM NAA，在此培养基上培养8周，73.4%的愈伤组织产生芽，每1cm 3愈伤组织产生芽3 ~ 6个。将不定芽切下并在补充有14.8 μM 3-吲哚丁酸的半强度Murashige和Skoog培养基中生根。培养10 d后生根，生根率为98%~ 100%。在遮荫温室的容器基质中生长的试管苗成活率超过95%。在过去的6年中，超过400万株再生使用这个既定的程序，并没有体细胞无性系变异。再生植株的千花鲜重、千花干重、CGA含量和干花产量与扦插繁殖的“金翠蕾”无显著差异，表明该方法再生植株稳定。这种已建立的体外培养方法已使这种药用植物在超过1500公顷的生产场地上实现了快速的商业化生产。
'Jincuilei' is a mutant selected from Lonicera macranthoides Hand.-Mazz. It produces abundant flowers that never open with a chlorogenic acid (CGA) content up to 6.0%. Propagation through rooting or grafting has only a 30% survival rate. This study was undertaken to establish an efficient protocol for rapidly regenerating this mutant. Leaf explants were inoculated on Gamborg's B 5 medium supplemented with different concentrations of 6-benzyladenine (BA) and 2,4-dichlorophenozyacetic acid (2,4-D). The optimal combination for callus induction was 4.4 μM BA with 2.26 μm 2,4-D, which resulted in 86.7% of leaf explants producing calluses in 4 weeks. Calluses produced from this optimal medium were cultured on B 5 medium containing different concentrations of kinetin (KT) and α-naphthalene acetic acid (NAA). The best formulation for shoot induction was B 5 medium containing 0.9 μM KT and 5.4 μM NAA in which 73.4% of cultured calluses produced shoots in 8 weeks, and shoot numbers ranged from three to six per callus piece (1 cm 3 ). Adventitious shoots were cut and rooted in half-strength Murashige and Skoog medium supplemented with 14.8 μM 3-indolebutyric acid. Roots initiated 10 d after culture, and rooting percentages ranged from 98% to 100%. Plantlets grown in a container substrate in a shaded greenhouse had over a 95% survival rate. During the last 6 years, over four million plantlets were regenerated using this established procedure, and there was no somaclonal variation. Fresh and dry weights of 1000 flowers, CGA contents, and dry flower yields of the regenerated plants were not significantly different from those of the stock 'Jincuilei' propagated by cutting, indicating that plants regenerated from this established procedure were stable. This established in vitro culture method has led to rapid commercial production of this medicinal plant on more than 1500 ha of production field.