关键词:小麦; 蛋白磷酸酶2A; 启动子; 顺式作用元件; GUS组织化学染色; GUS定量分析 Cloning and Expression Analysis of Protein Phosphatase 2A Gene TaPP2AbB″-α Promoter in Wheat YI Heng1,2, LI Ang2, LIU Hui-Min1, JING Rui-Lian2,* 1 College of Bioengineering, Shanxi University, Taiyuan 030006, China
2 National Key Facility for Crop Gene Resources and Genetic Improvement / Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing 100081, China
Fund:This study was supported by the National Natural Science Foundation of China (31201206) and the Agricultural Science and Technology Innovation Program (CAAS) AbstractProtein phosphatase 2A (PP2A) is a heterotrimeric protein, consisting of a scaffolding subunit (A), a catalytic subunit (C), and a member of four families of regulatory subunits (B). PP2A plays significant roles in the pathway responding to abiotic stresses in plants. TaPP2AbB″-α, a member of regulatory subunit B″ in wheat ( Triticum aestivum L.), enhanced root development and could develop more lateral roots in the gene overexpressed Arabidopsis, especially under the osmotic stresses of mannitol and NaCl. In order to elucidate transcriptional regulatory mechanism of the promoter PB″α of TaPP2AbB″-α, we isolated an 1899 bp full-length sequence of promoter PB″α from a drought-tolerant wheat cultivar Hanxuan 10. The PB″α sequence contained TATA-box, CAAT-box and a series of cis-acting elements responding to drought and osmotic stresses, such as elements of EECCRCAH1 (from -1058 to -1052 bp), GCCCORE (from -1073 to -1068 bp) and MYCCONSE (from -1179 to -1174 bp). The full-length promoter PB″α and five 5′-end truncated PB″α promoters in different lengths fused with the reporter gene β-glucuronidas ( GUS) were transformed into Arabidopsis, respectively. The histochemical staining results showed that the full-length promoter PB″α, and the deletion fragments of PB″α-1545 and PB″α-1389 could drive GUS gene expression in shoots and roots of transgenic Arabidopsis seedlings. As the result of quantitative fluorometric GUS assay, only the expression of PB″α, PB″α-1545 and PB″α-1389 could be up-regulated by salt and osmotic stresses in transgenic Arabidopsis lines, and the active region of PB″α promoter was located in the interval between -1389 bp and -946 bp. In conclusion, PB″α has strong basic promoter activity which is up-regulated significantly by salt and osmotic stresses. These findings contribute to the selection of a suitable promoter for crop improvement.
Keyword:Wheat; PP2A; Promoter; cis-acting Element; GUS histochemical staining; Quantitative fluorometric GUS assay Show Figures Show Figures
图8 胁迫条件下转基因拟南芥GUS定量分析 * 和* * 分别表示胁迫处理与对照(CK)在0.05和0.01概率水平的差异显著(t检验)。Fig. 8 Quantification of GUS activity in transgenic Arabidopsis under different stresses * and * * indicate significant difference between stress treatment and the control (CK) at the 0.05 and 0.01 probability levels, respectively (t-test).
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