Adventitious shoot regeneration of oriental lily (Lilium orientalis) and genetic stability evaluation based on ISSR marker variation
Adventitious shoot regeneration of oriental lily (Lilium orientalis) and genetic stability evaluation based on ISSR marker variation
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DOI：
10.1007/s11627-012-9429-0
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发表时间：
2012-04-01
影响因子：
2.6
通讯作者：
Yang, Guochen
中科院分区：
文献类型：
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作者：
Liu, Xiaomei;Yang, Guochen
以东方百合（Lilium orientalis）为试材，建立了以鳞片和叶片为外植体的再生体系。以离体培养的叶片为外植体，在含有噻苯隆（TDZ）或6-苄氨基嘌呤（BA）和萘乙酸（NAA）的Murashige和Skoog培养基上诱导出不定芽。叶片在TDZ 10.8 μ M + NAA 0.54 μ M的培养基上再生率最高，达74.2%。鳞茎鳞片外植体在含0.54 μ M TDZ和0.54 μ M NAA的培养基上产生的芽平均数最高，为4.4。不定芽生根率为79.2%~100%。生根苗经温室驯化后成活。卡那霉素浓度对不定芽再生的影响也进行了评价，100毫克升（-1）的值被建议作为百合转化的致死剂量。利用18个ISSR标记对再生苗进行遗传稳定性分析。11个引物共扩增出70条清晰、重复性好的条带。无性系衍生物和母株之间的遗传相似性指数在0.92~1.0之间。所有15个微繁殖后代和母株可以聚在一起，在一个主要的聚类相似性水平为92%。整个植株的体细胞无性系变异率估计为4.2%，表明直接从外植体再生芽的形成是一个安全的方法繁殖的“真正的类型”的植物。
A regeneration system was developed for oriental lily (Lilium orientalis) based on both leaf and bulb scale. Adventitious shoots were regenerated from leaves of in vitro cultures on Murashige and Skoog medium containing thidiazuron (TDZ) or 6-benzylaminopurine (BA) and naphthaleneacetic acid (NAA). The highest percent regeneration from leaf explants was 74.2%, being observed on medium containing 10.8 mu M TDZ and 0.54 mu M NAA. The highest mean number of shoots generated was 4.4 and was obtained from bulb scale explants on medium containing 0.54 mu M TDZ and 0.54 mu M NAA. Adventitious shoots were successfully rooted at rates ranging from 79.2% to 100%. The rooted plantlets survived after acclimatization in the greenhouse. The effect of kanamycin concentration on adventitious shoot regeneration was also evaluated, a value of 100 mg l(-1) being suggested as a lethal dose for lily transformation. Eighteen ISSR markers were employed to determine the genetic stability of the regenerated shoots in comparison to their mother plant. Eleven primers in total produced 70 clear and reproducible bands. Genetic similarity indicators among the clonal derivatives and the mother plant ranged from 0.92 to 1.0. All 15 micropropagated progenies and the mother plant could be grouped together in one major cluster with a similarity level of 92%. The somaclonal variation rate across the plantlets was estimated as 4.2%, indicating that direct shoot formation from explant regeneration is a safe method for multiplication of "true-to-type" plants.