Validation of Reference Genes for Gene Expression by Quantitative Real-Time RT-PCR in Stem Segments Spanning Primary to Secondary Growth in Populus tomentosa.
Validation of Reference Genes for Gene Expression by Quantitative Real-Time RT-PCR in Stem Segments Spanning Primary to Secondary Growth in Populus tomentosa.
复制标题
通过定量实时 RT-PCR 在毛白杨初生到次生生长的茎段中验证基因表达的参考基因
DOI：
10.1371/journal.pone.0157370
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发表时间：
2016
期刊：
影响因子：
3.7
通讯作者：
Wang H
中科院分区：
文献类型：
--
作者：
Wang Y;Chen Y;Ding L;Zhang J;Wei J;Wang H
杨树茎段垂直切段是分析木材形成过程中初生和次生生长相关基因表达模式的理想实验系统。合适的内部对照基因对于基因表达的定量真实的时间PCR（qRT-PCR）测定是必不可少的。本研究以毛白杨垂直茎段为材料，对8个候选参考基因的表达稳定性进行了研究。通过软件包geNorm、Normandy和BestKeeper的分析表明，在毛白杨茎的发育阶段，基因核糖体蛋白（RP）和微管蛋白β（TUBB）是最不稳定的，并且三个参考基因的组合，真核翻译起始因子5A（eIF 5A），肌动蛋白（ACT 6）和延伸因子1-β（EF 1-β）可以为毛白杨经历初生和次生生长的茎段中靶基因表达的qRT-PCR分析提供准确和可靠的标准化。这些结果为毛白杨茎段的转录分析提供了重要信息，有助于提高毛白杨垂直茎段基因表达数据的质量，为研究木材形成提供了良好的植物系统。
The vertical segments of Populus stems are an ideal experimental system for analyzing the gene expression patterns involved in primary and secondary growth during wood formation. Suitable internal control genes are indispensable to quantitative real time PCR (qRT-PCR) assays of gene expression. In this study, the expression stability of eight candidate reference genes was evaluated in a series of vertical stem segments of Populus tomentosa. Analysis through software packages geNorm, NormFinder and BestKeeper showed that genes ribosomal protein (RP) and tubulin beta (TUBB) were the most unstable across the developmental stages of P. tomentosa stems, and the combination of the three reference genes, eukaryotic translation initiation factor 5A (eIF5A), Actin (ACT6) and elongation factor 1-beta (EF1-beta) can provide accurate and reliable normalization of qRT–PCR analysis for target gene expression in stem segments undergoing primary and secondary growth in P. tomentosa. These results provide crucial information for transcriptional analysis in the P. tomentosa stem, which may help to improve the quality of gene expression data in these vertical stem segments, which constitute an excellent plant system for the study of wood formation.