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首页> 外文期刊>Journal of Experimental Botany >Expression profile analysis of genes involved in cell wall regeneration during protoplast culture in cotton by suppression subtractive hybridization and macroarray
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Expression profile analysis of genes involved in cell wall regeneration during protoplast culture in cotton by suppression subtractive hybridization and macroarray

机译：抑制性消减杂交和宏阵列技术分析棉花原生质体培养过程中参与细胞壁再生的基因的表达谱分析

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摘要

The molecular mechanisms underlying cell wall biosynthesis are poorly understood. In this study, microscopic analysis showed that protoplasts generated a new cell wall within 48 h after transfer to a wall-regeneration medium. To identify genes related to cell wall biosynthesis in cotton, suppression subtractive hybridization was used to visualize differential gene expression at seven time points within the first 48 h. In total, 412 differentially expressed sequence tags (ESTs; 3-fold) were identified, and 210 unigenes were sequenced successfully. As confirmed by reverse-transcription PCR (RT-PCR) and real-time quantitative reverse-transcription PCR (QRT-PCR) analysis, the selected genes displayed complex expression patterns during cell wall regeneration from protoplasts and included most previously published cell-wall-associated genes. ESTs similar to cell-wall-protein genes, such as proline-rich protein (PRPL), glycine-rich protein (GRP), extension (EPR1), fasciclin-like arabinogalactan protein (FLA2), and expensing-like protein (EXLA and EXLB), which might participate in primary cell wall or secondary cell wall construction and modification, were up-regulated during cell wall regeneration from protoplasts. Sucrose synthase, an important enzyme in the sugar signalling pathway, played important roles in cellulose biosynthesis. Our findings also highlighted the function of some transcription factors during cell wall regeneration from protoplasts, including the squamosa promoter binding protein-like 14 (SPL14), NAC, Gbiaa-re, MYB, WRKY, swellmap 1 (SMP1), RAD5, and zinc finger family protein, as well as the enrichment of Ca2+-calmodulin signal molecules. On the basis of the gene expression profiles, a model of cell wall regeneration from protoplasts derived from cotton suspension cultures is proposed.
机译：细胞壁生物合成的分子机制了解甚少。在这项研究中，显微镜分析显示，原生质体转移到壁再生培养基后48小时内产生了新的细胞壁。为了鉴定与棉花中细胞壁生物合成相关的基因，使用抑制消减杂交技术在前48小时内的七个时间点观察差异基因表达。总共鉴定了412个差异表达的序列标签（EST；> 3倍），并成功测序了210个单基因。正如逆转录PCR（RT-PCR）和实时定量逆转录PCR（QRT-PCR）分析所证实的那样，所选基因在原生质体细胞壁再生过程中显示出复杂的表达模式，其中包括大多数先前发表的细胞壁相关基因。与细胞壁蛋白基因相似的EST，例如富含脯氨酸的蛋白（PRPL），富含甘氨酸的蛋白（GRP），延伸（EPR1），类fasciclin的阿拉伯半乳聚糖蛋白（FLA2）和类似消耗性蛋白（EXLA和可能参与原代细胞壁或次生细胞壁的构建和修饰的EXLB）在原生质体细胞壁再生过程中被上调。蔗糖合酶是糖信号途径中的重要酶，在纤维素生物合成中起着重要作用。我们的发现还强调了原生质体细胞壁再生过程中某些转录因子的功能，包括鳞状启动子结合蛋白样14（SPL14），NAC，Gbiaa-re，MYB，WRKY，swellmap 1（SMP1），RAD5和锌手指家族蛋白，以及Ca 2 + -钙调蛋白信号分子的富集。基于基因表达谱，提出了从棉花悬浮培养物中原生质体再生细胞壁的模型。

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