Generation of transgenic wheat (Triticum aestivum L.) accumulating heterologous endo-xylanase or ferulic acid esterase in the endosperm
Generation of transgenic wheat (Triticum aestivum L.) accumulating heterologous endo-xylanase or ferulic acid esterase in the endosperm
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DOI：
10.1111/j.1467-7652.2009.00490.x
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发表时间：
2010-04-01
影响因子：
13.8
通讯作者：
Scheller, Henrik V.
中科院分区：
文献类型：
--
作者：
Harholt, Jesper;Bach, Inga C.;Scheller, Henrik V.
内切木聚糖酶（来自枯草芽孢杆菌）或阿魏酸酯酶（来自尼日尔曲霉）在胚乳特异性1DX 5谷蛋白启动子的控制下在小麦中表达。使用具有和不具有内质网滞留信号（Lys-Asp-Glu-Leu）KDEL的构建体。在所有四种情况下都回收了转基因植物，但无论是否添加KDEL，都没有观察到定性差异。转基因谷物中的内切木聚糖酶活性相对于野生型增加了两到三倍。籽粒干瘪，质量减少25%-33%。细胞壁的广泛分析显示阿拉伯糖与木糖的比率增加10%-15%，水可提取的阿拉伯木聚糖的比例增加50%，并且水可提取的阿拉伯木聚糖的MW从主要大于85 kD转变为2 - 85 kD。表达阿魏酸酯酶的籽粒也皱缩，种子重量减少20%-50%。在野生型谷粒中没有检测到阿魏酸酯酶活性，而在转基因株系中检测到阿魏酸酯酶活性。籽粒细胞壁中水不可提取的阿拉伯木聚糖增加15%-40%，单体阿魏酸减少13%-34%。在所有植物中，观察到的变化与植物响应一致，植物响应通过增加阿拉伯木聚糖生物合成和交联来最小化异源表达酶的影响。
P>Endo-xylanase (from Bacillus subtilis) or ferulic acid esterase (from Aspergillus niger) were expressed in wheat under the control of the endosperm-specific 1DX5 glutenin promoter. Constructs both with and without the endoplasmic reticulum retention signal (Lys-Asp-Glu-Leu) KDEL were used. Transgenic plants were recovered in all four cases but no qualitative differences could be observed whether KDEL was added or not. Endo-xylanase activity in transgenic grains was increased between two and threefold relative to wild type. The grains were shrivelled and had a 25%-33% decrease in mass. Extensive analysis of the cell walls showed a 10%-15% increase in arabinose to xylose ratio, a 50% increase in the proportion of water-extractable arabinoxylan, and a shift in the MW of the water-extractable arabinoxylan from being mainly larger than 85 kD to being between 2 and 85 kD. Ferulic acid esterase-expressing grains were also shrivelled, and the seed weight was decreased by 20%-50%. No ferulic acid esterase activity could be detected in wild-type grains whereas ferulic acid esterase activity was detected in transgenic lines. The grain cell walls had 15%-40% increase in water-unextractable arabinoxylan and a decrease in monomeric ferulic acid between 13% and 34%. In all the plants, the observed changes are consistent with a plant response that serves to minimize the effect of the heterologously expressed enzymes by increasing arabinoxylan biosynthesis and cross-linking.