Abstract In vitro selection of Withania somnifera has been incurred against Alternaria alternata by selection pressure of fungal toxin in an innovative way. Solid green callus was maintained in Murashige and Skoog's medium supplemented with 6-Benzylaminopurine (2 mgL−1). Toxin was partially purified from A. alternata broth culture. Elite clone was generated by gradual intoxication of solid green callus in the toxin containing medium (5–400 mgL−1). Three highest dose of toxin tolerance was selected viz. 50 mgL−1, 100 mgL−1 and 200 mgL−1. Plants were successfully regenerated from those toxin selected callus. Eventually, various defense enzymes and total phenolics were measured in regenerated plants and toxin selected callus. Callus selected from 100 mgL−1 toxin dose and plants obtained from them give the best results among all and showed more than 2–5 fold increase of defense enzyme activities and total phenolics compared to control. Furthermore, those plants when challenge inoculated showed significant reduction (more than 70%) of leaf spot incidence. Augmentation of disease resistance persists for at least two successive generations. Furthermore, the increase in defense molecules also correlated with increased nitric oxide generation (73%). Presence of withaferin A and withanolide A has also been confirmed in the toxin regenerated plants by High-performance thin-layer chromatography (HPTLC) analysis. Over all observations suggests that fungal toxin at a concentration 100 mgL−1 and by habituation technique can be use as a selection method for the production of disease resistant medicinally important crop like W. somnifera without altering its secondary metabolite production.

中文翻译：

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Withania somnifera 优良克隆抗链格孢引起的叶枯病的体外筛选

摘要 通过真菌毒素的选择压力，以创新的方式对催眠睡茄进行了体外选择以对抗链格孢。实心绿色愈伤组织保持在补充有 6-苄氨基嘌呤 (2 mgL-1) 的 Murashige 和 Skoog 培养基中。从 A. alternata 肉汤培养物中部分纯化毒素。通过在含有毒素的培养基 (5–400 mgL-1) 中逐渐中毒固体绿色愈伤组织产生精英克隆。选择了三种最高剂量的毒素耐受性，即。50 mgL-1、100 mgL-1 和 200 mgL-1。从那些毒素选择的愈伤组织中成功地再生出植物。最后，在再生植物和毒素选择的愈伤组织中测量了各种防御酶和总酚类物质。从 100 mgL-1 毒素剂量中选择的愈伤组织和从中获得的植物给出了最好的结果，与对照相比，防御酶活性和总酚类物质增加了 2-5 倍以上。此外，这些植物在接种攻击时表现出叶斑发生率的显着降低（超过 70%）。抗病性的增强持续至少连续两代。此外，防御分子的增加也与一氧化氮生成的增加（73%）相关。通过高效薄层色谱 (HPTLC) 分析也证实了在毒素再生植物中存在 withaferin A 和 withanolide A。