Characterization of expression of the OsPUL gene encoding a pullulanase-type debranching enzyme during seed development and germination in rice
Characterization of expression of the OsPUL gene encoding a pullulanase-type debranching enzyme during seed development and germination in rice
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水稻种子发育和发芽过程中编码普鲁兰酶型脱支酶的 OsPUL 基因的表达特征
DOI：
10.1016/j.plaphy.2009.02.001
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发表时间：
2009-05-01
影响因子：
6.5
通讯作者：
Liu, Qiao-Quan
中科院分区：
文献类型：
--
作者：
Li, Qian-Feng;Zhang, Gui-Yun;Liu, Qiao-Quan
淀粉脱支酶（DBEs）是谷物淀粉代谢的关键酶，具有淀粉合成和降解的双重功能。然而，它们在这一途径中的确切作用，特别是它们的表达谱，仍不清楚。在本研究中，我们采用实时荧光定量PCR （Q-PCR）方法对编码普鲁兰酶型DBE的OsPUL基因在不同组织和不同发育阶段种子中的表达模式进行了分析。结果表明，该基因仅在种子中表达。此外，克隆了1177 bp的OsPUL启动子序列，发现该区域存在GCN4、AACA等胚乳特异性基序。然后将该启动子与GUS报告基因融合，并在转基因水稻中仔细研究了其表达。组织化学和荧光分析的数据表明，OsPUL启动子能够驱动靶基因具有高水平的胚乳特异性表达。OsPUL基因在种子发育的整个过程中都保持较高的表达水平，在中后期达到高峰。这与Q-PCR内源转录分析结果非常一致。此外，种子萌发试验表明，OsPUL启动子在种子萌发后期发挥积极作用。在整个灌浆期和种子萌发后期，OsPUL基因的表达维持在显著水平，这与其参与淀粉合成代谢和分解代谢相一致。(C) 2009 Elsevier Masson SAS。版权所有。
Starch-debranching enzymes (DBEs) are key enzymes involved in starch metabolism in cereals, having a dual function, in both starch synthesis and degradation. However, their precise roles in this pathway, particularly their expression profiles, remain unclear. In the present study, we performed a quantitative real-time PCR (Q-PCR) analysis of the expression pattern of the OsPUL gene encoding a pullulanase-type DBE in different tissues as well as in seeds at different developmental stages. The results showed that this gene was expressed only in seeds. In addition, the 1177-bp OsPUL promoter sequence was cloned, and some endosperm-specific motifs such as the GCN4 and AACA motifs were observed to exist in this region. The promoter Was then fused with the GUS reporter gene and its expression was carefully investigated in transgenic rice. The data from both histochemical and fluorometric analyses showed that the OsPUL promoter Was capable of driving the target gene to have a high level of endosperm-specific expression. The OsPUL gene maintained a relatively high expression level during the entire period of seed development, and peaked in the middle and late stages. This observation Was Very consistent with that of the endogenous transcription analysis by Q-PCR. Furthermore, the seed germination experiment showed that the OsPUL promoter actively functions in the late stage of seed germination. The expression of the OsPUL gene was maintained at a significant level during the entire grain filling period and in the late stage of seed germination, which coincided with its involvement in starch anabolism and catabolism. (C) 2009 Elsevier Masson SAS. All rights reserved.