Plant Regeneration from petiole Callus of Amorphophallus albus and Analysis of Somaclonal Variation of Regenerated Plants by RAPD and ISSR Markers

本文概述了一種用於中國藥用植物白魔芋組織培養的簡便方法，即從葉柄愈傷組織誘導植株再生。採用2個月苗齡植株的幼葉柄為外植體，接種於附加5.37μMNAA和4.44μMBAMS培養基，愈傷組織誘導頻率高達76.4±3.2%。所誘導的愈傷組織類型各異，其中Ⅲ型愈傷組織用於形態發生的建立。Ⅲ型愈傷組織培養於加有適宜濃度的NAA和BA或KT組合的MS培養基上可形成擬球莖結構，繼續培養擬球莖結構便可產生芽和根。最佳形態發生反應出現在細胞分裂素與生長素濃度比約為4:1的培養基上，即擬球莖形成頻率約為70%，每塊愈傷組織長可形成6~8個擬球莖。將擬球莖繼代培養於其形成的培養基上，不另需生根培養就可獲得具有完整根系的植株。具有根系的再生植株移栽至大田後的存活率在90%以上。採用兩種分子標記技術，即隨機擴增多態性(RAPD)和簡單重複間序列(ISSR)，對20株再生植株進行變異檢測並發現植株中存在變異。在這20株植株中，RAPD和ISSR分別檢測到20.8%和39.0%多態性頻率。對RAPD和ISSR資料分別進行聚類分析，結果表明21株植株（20株再生植株和1株母本）的遺傳相似係數分別為0.973和0.917，且能分為不同的族群。白魔芋組織培養中所誘導的高頻無性系變異，可能有助於有用突變體的篩選從而用於其遺傳改良。

關鍵字

白魔芋；器官發生；分子標記；無性系變異

並列摘要

A simple procedure has been outlined for plant regeneration of Amorphophallus albus Liu & Wei, a native medicinal plant in China, from petiole-derived callus. Calli were induced at a high frequency of 76.4±3.2% from petiole explants excised from two-month-old plants on Murashige and Skoog (MS) medium supplemented with 5.37 μM α-naphthaleneacetic acid (NAA) and 4.44 μM 6-benzyladenine (BA). Of the different types of callus induced, type III callus was selected for morphogenesis induction. Culture of the callus on MS medium containing proper NAA and BA or KT combinations resulted in formation of corm-like structure (CLS) that produced shoots and roots during further culture. The optimal morphogenetic response was observed on the media with a cytokinin/auxin ratio of about 4:1, which resulted in more than 70% CLS formation and 6~8 CLSs per callus. Complete plantlets with well-developed root systems were obtained from these CLSs by subculturing them on the original media from which they had been derived without a separate rooting culture. Transfer of the plantlets with roots to soil resulted in a more than 90% survival rate. Analysis of 20 regenerated plants by two molecular markers, randomly amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR), revealed somaclonal variation in the regenerated plants. The percentage of polymorphic bands in RAPD and ISSR analysis were respectively 20.8% and 39.0% for the 20 plants. Cluster analysis indicated that the genetic similarity values calculated on the basis of RAPD and ISSR data among the 21 plants (20 regenerated and one donor plant) were, respectively, 0.973 and 0.917, which allowed classification of the plants into distinct groups. A high-frequency somaclonal variation induced in A. albus tissue culture may help in the selection of useful variants that may be induced to improve this important corp.

並列關鍵字

Amorphophallus albus Liu & Wei ； Organogenesis ； Molecular marker ； Somaclonal variation

國際替代計量

Plant Regeneration from petiole Callus of Amorphophallus albus and Analysis of Somaclonal Variation of Regenerated Plants by RAPD and ISSR Markers

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