水稻α-淀粉酶基因的表达模式与颖花开放的关系

doi:10.3864/j.issn.0578-1752.2023.07.006

摘要/Abstract

摘要：

【目的】淀粉降解与水稻浆片膨大和颖花开放过程密切相关，探究α-淀粉酶基因在颖花开放过程中的作用，为杂交水稻制种效率及产量的提高提供理论依据。【方法】在水稻扬花时，利用稀释碱性品红溶液进行离体穗子吸水试验，观察碱性品红在颖花中残留的组织，通过碘-碘化钾染色法确定11—14期（依据雄蕊发育分期）淀粉粒的分布变化，并通过RT-PCR、RT-qPCR和GUS报告基因检测多个α-淀粉酶基因在此期间的时空表达模式。【结果】水稻颖花开放前，内外稃片通过相互嵌合的钩合槽（marginal tissues of palea，mtp）将浆片和雌雄蕊封闭在内。当颖花开放时，浆片快速膨大，使得内外稃片的钩合点松开。扬花期间，离体穗子在稀释碱性品红溶液中吸水后，碱性品红染料主要残留在内外稃片钩合槽和浆片相连处组织以及花丝中。碘染试验显示，在12期（颖花开放前），淀粉粒主要分布在雄蕊和内外稃片钩合槽，浆片中也有少量淀粉粒，在13—14期（颖花开放中），内外稃片钩合槽和浆片中的淀粉粒均降解。RT-PCR分析发现OsRAmy2A和OsRAmy3D的表达量从12期开始上升，至13—14期表达量显著增强，到受精后1 d（1 day after pollination，DAP1）表达量又明显下降，OsRAmy3E和OsRAmy3F在此过程中持续表达，OsRAmy3F表达量弱于OsRAmy3E。RT-qPCR分析显示，在11—14期，OsRAmy2A表达量变化最显著，其次是OsRAmy3A和OsRAmy3E，OsRAmy3F的表达量变化幅度最不明显，OsRAmy2A和OsRAmy3A在13—14期表达量显著增加，而OsRAmy3E和OsRAmy3F在不同时期均表达，在13—14期表达量略有升高。在12期，OsRAmy2A主要在内外稃以及内外稃片钩合槽上表达，在13—14期主要在内外稃片钩合槽、浆片以及花丝上表达。【结论】水稻颖花开放过程中淀粉粒在mtp和浆片中明显降解，与OsRAmy2A、OsRAmy3D等α-淀粉酶基因时空表达模式相对应，可能与水稻浆片膨大导致颖花开放过程密切相关。

关键词: 水稻, α-淀粉酶, 颖花开放, 淀粉粒, 浆片膨大

Abstract:

【Objective】 Starch degradation is involved in lodicule absorbing abundant water and swelling during rice floret opening, but the amylase genes associated with this process have not been identified yet. 【Method】 To identify the swelling of tissues during floret opening, the in vitro rice panicles absorbed diluted Fuchsin basic and the dye remains were observed after florets were closing again. The starch grain distribution in rice florets before and during anthesis from stage 11 to stage 14 (according to 14 stages of rice anther development) was detected using iodide staining. The spatial-temporal expression patterns of 10 α-amylase genes were detected by RT-PCR, RT-qPCR and GUS staining. 【Result】 Before floret opening, the stamens, pistils and lodicules are enclosed by the lemma and palea through marginal tissues of palea (mtp). Rapid swelling of the lodicules causes floret opening by separating the lemma from the palea. After the in vitro panicles absorbed diluted Fuchsin basic during floret opening, the dye remains were observed located in the joint between mtp and lodicules and filaments. Iodide staining showed that the starch grains were mainly located in the stamens and mtp and a small amount of starch grains in the lodicules at stage 12 (before floret opening), whereas the starch grains in the mtp and lodicules were almost completely degraded at stage 13-14 (during floret opening). RT-PCR showed that OsRAmy2A and OsRAmy3D began to express from stage 12 and were expressed with high levels at stage 13-14. The expression levels of the two genes decreased at DAP1 (1 day after pollination). OsRAmy3E and OsRAmy3F kept expressed during this process. The expression level of OsRAmy3E was higher than that of OsRAmy3F. The RT-qPCR analysis showed that the expression level of OsRAmy2A increased most dramatically at stage 13-14, followed by OsRAMy3A and OsRAMy3E. Further, the transgenic plants expressing the GUS reporter gene driven by the OsRAmy2A promoter were generated. The GUS signaling was located only in the lemma, palea and mtp at stage 12 and the expression of the GUS gene driven by the RAmy2A promoter was induced in the mtp, lodicules and filaments at stage 13-14. 【Conclusion】 These data indicated that starch grain degradation in the mtp and lodicules at stage 13-14 might be related with high expression levels of some α-amylase genes such as OsRAmy2A and OsRAmy3D, probably involved in controlling lodicules swelling and floret opening in rice.

Key words: rice, α-amylase, floret opening, starch grain, lodicules swelling

张绩, 周上铃, 何发, 刘莉莎, 张玉娟, 何晋宇, 杜晓秋. 水稻α-淀粉酶基因的表达模式与颖花开放的关系[J]. 中国农业科学, 2023, 56(7): 1275-1282.

ZHANG Ji, ZHOU ShangLing, HE Fa, LIU LiSha, ZHANG YuJuan, HE JinYu, DU XiaoQiu. Expression Pattern of the Rice α-Amylase Genes Related with the Process of Floret Opening[J]. Scientia Agricultura Sinica, 2023, 56(7): 1275-1282.

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