Protoplast isolation and culture from carob ( Ceratonia siliqua ) hypocotyls: ability of regenerated protoplasts to produce mannose-containing polysaccharides
Protoplast isolation and culture from carob ( Ceratonia siliqua ) hypocotyls: ability of regenerated protoplasts to produce mannose-containing polysaccharides
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角豆（Ceratonia siliqua）下胚轴原生质体分离和培养：再生原生质体产生含甘露糖多糖的能力
DOI：
10.1111/j.1399-3054.2007.00878.x
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发表时间：
2007
影响因子：
6.4
通讯作者：
Sotiriou P
中科院分区：
文献类型：
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作者：
Sotiriou P
以角豆（Ceratonia siliquaL.）胚胎组织具有再生细胞壁、分裂和合成半乳甘露聚糖（一种有价值的工业多糖）的能力。从角豆下胚轴钩分离的原生质体在24小时内再生细胞壁。在培养2天后观察到再生细胞的第一次分裂。当幼苗在漫射光下生长时，成功分裂的百分比最高，下胚轴钩在原生质体分离溶液中孵育之前被原生质体化1小时，并且原生质体在漫射光下培养。培养9天后，产生了由8个细胞组成的细胞簇，其经历进一步的有丝分裂，并预期导致愈伤组织形成。通过用外源d-[U-14 C]葡萄糖、d-[U-14 C]甘露糖和d-[2 - 3 H]甘露糖进行放射性标记，研究了原生质体再生过程中多糖和寡糖的合成，这些标记分别产生均匀、中等特异性和高度特异性标记。正如细胞壁单糖的放射性分布所揭示的那样，再生体沉积了新的壁聚合物，这些聚合物与分离出原生质体的下胚轴所合成的聚合物明显不同。再生剂沉积了大量的胼胝质和少量的含半乳糖、阿拉伯糖和甘露糖的聚合物。后者包括葡糖醛酸甘露聚糖，如通过涉及部分酸水解然后是β-葡糖醛酸酶（EC 3.2.1.31）消化的新方法所证明的。再生的原生质体还释放可溶性胞外碳水化合物：多糖，似乎主要是酸性阿拉伯半乳聚糖，和寡糖，主要是中性的，含有葡萄糖，半乳糖和甘露糖。我们得出结论，再生角豆原生质体是一个有用的系统，用于研究碳水化合物的分泌，包括富含甘露糖的多糖和寡糖。
The aim of this study was to isolate protoplasts from carob (Ceratonia siliquaL.) embryonic tissues with the ability to regenerate cell walls, divide and synthesize galactomannan, a valuable polysaccharide for industry. Protoplasts isolated from carob hypocotyl hooks regenerated cell walls within 24 h. The first divisions of the regenerated cells were observed after 2 days of culture. The highest percentage that successfully divided was achieved when the seedlings were grown under diffuse light, the hypocotyl hooks were plasmolysed for 1 h before incubation in the protoplast isolation solution and the protoplasts were cultured under diffuse light. After 9 days of culture, cell clusters, consisting of eight cells, had been produced, which underwent further mitotic divisions and which were expected to lead to callus formation. Polysaccharide and oligosaccharide synthesis during protoplast regeneration was studied by radiolabelling with exogenousd‐[U‐14C]glucose,d‐[U‐14C]mannose ord‐[2‐3H]mannose, which gave rise to uniform, moderately specific and highly specific labelling, respectively. As revealed by the radioactivity distribution in cell wall monosaccharides, the regenerants deposited new wall polymers that differed markedly from those being synthesized by the hypocotyls from which the protoplasts had been isolated. The regenerants deposited large amounts of callose and smaller amounts of galactose‐, arabinose‐ and mannose‐containing polymers. The latter included glucuronomannan, as demonstrated by a new method involving partial acid hydrolysis followed by β‐glucuronidase (EC 3.2.1.31) digestion. The regenerating protoplasts also released soluble extracellular carbohydrates: polysaccharides which appeared to be mainly acidic arabinogalactans, and oligosaccharides which were mainly neutral and contained glucose, galactose and mannose. We conclude that regenerating carob protoplasts are a useful system for studying carbohydrate secretion, including mannose‐rich poly‐ and oligosaccharides.