Key message

An efficient, high-frequency, and robust in vitro regeneration protocol was developed using cotyledon and hypocotyl explants from mulberry (Morus indica cv. G4) seedlings.

Abstract

Mulberry (Morus) is a perennial tree species with a wide range of commercial applications. Its leaves are predominantly used for feeding the monophagous silkworm (Bombyx mori L.) globally. In this study, the effects of plant growth regulators, additives and elevated levels of macronutrients, on in vitro adventitious shoot induction, shoot elongation, and rooting, were investigated. It was found that modified Murashige and Skoog (MS) medium supplemented with thidiazuron (0.5 mg/L) provided the most suitable medium for adventitious shoot bud induction, with a regeneration frequency of 88.62%, and yielded adventitious shoot buds of 10.60 ± 0.30 per cotyledon explant. Additionally, the MS medium fortified with 6-benzylaminopurine (1.0 mg/L), gibberellic acid (1.5 mg/L)_, silver nitrate (2 mg/L), putrescine dihydrochloride (1 mg/L), activated charcoal (AC, 0.2%), and supplementary dosage of calcium chloride (515 mg/L) resulted in the highest frequency of shoot elongation, spontaneous root induction, and the longest shoot length. In this medium, no hyperhydricity of regenerated shoots/leaves was observed. We observed the longest adventitious root length and secondary root length of the shoots grown on MS medium supplemented with indole-3-butyric acid (2 mg/L) and AC (0.2%). The frequencies of ex vitro survival of plantlets after hardening were 90–95% and 95–100%, under laboratory and field-like conditions, respectively. Even though in vitro regeneration protocol in mulberry is genotype-dependent and explant-specific, the robust regeneration protocol developed in this study could find its applications in genome editing and genetic transformation using cotyledon and hypocotyl explants of other cultivars.

中文翻译：

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从桑树 cv. 的幼苗外植体中体外再生桑树植物。G4 通过直接器官发生

关键信息

使用来自桑树 ( Morus indica cv. G4) 幼苗的子叶和下胚轴外植体开发了一种高效、高频和稳健的体外再生方案。

抽象的

桑树（Morus）是一种多年生树种，具有广泛的商业用途。它的叶子主要用于喂养全球的单食蚕（Bombyx mori L.）。在这项研究中，研究了植物生长调节剂、添加剂和大量营养素水平升高对体外不定芽诱导、芽伸长和生根的影响。结果发现，添加噻唑隆（0.5 mg/L）的改良 Murashige 和 Skoog (MS) 培养基为不定芽诱导提供了最合适的培养基，再生频率为 88.62%，产生的不定芽芽为 10.60 ± 0.30 个/子叶外植体。此外，MS 培养基强化了 6-苄氨基嘌呤 (1.0 mg/L)、赤霉酸 (1.5 mg/L) _、硝酸银（2mg/L）、二盐酸盐腐胺（1mg/L）、活性炭（AC，0.2%）和氯化钙的补充剂量（515mg/L）导致枝条伸长频率最高，自发根诱导，和最长的芽长。在该培养基中，未观察到再生芽/叶的超水合。我们观察到在补充有 indole-3-butyric 酸 (2 mg/L) 和 AC (0.2%) 的 MS 培养基上生长的芽的最长不定根长度和次生根长度。体外频率在实验室和类似田间条件下，硬化后幼苗的存活率分别为 90-95% 和 95-100%。尽管桑树的体外再生方案是基因型依赖性和外植体特异性的，但本研究中开发的强大再生方案可以在使用其他品种的子叶和下胚轴外植体的基因组编辑和遗传转化中找到其应用。