Utilization of Heterologous Expression of Hvsusiba2 to Increase Grain and Stalk Starch Contents and Amylopectin Synthase Activity in Rice
-
摘要: Hvsusiba2是一个来源于大麦的糖信号转录因子，它作用于淀粉合成途径的上游，是大麦淀粉合成途径的关键调控因子。本研究利用前期研究获得的转Hvsusiba2水稻纯合株系，探讨Hvsusiba2转录因子过表达对水稻淀粉合成的影响，并通过酶谱分析方法分析Hvsusiba2在水稻中的异源表达对支链淀粉合成同工酶的调控作用。结果显示：Hvsusiba2在水稻中的异源表达能够提高籽粒中总淀粉含量，增加淀粉含量达4.33%~9.22%，茎鞘的淀粉含量提高1倍，幼嫩种子发育过程中淀粉累积快速增加期为花后15 d起。对该时期的支链淀粉合成酶同工酶分析提示异源表达Hvsusiba2是通过提高籽粒中淀粉分支酶中淀粉分支酶SBEIIb、淀粉分支酶SBEI、异淀粉酶Isoamylase以及淀粉酶Amylase的活性改变水稻的支链淀粉合成，但淀粉分支酶中SBEIIa的活性不受Hvsusiba2转录因子的调控。Abstract: Hvsusiba2 is a transcription factor acting on the upstream of starch synthesis pathway. It is the key starch synthesis regulator in barley. To understand whether the gene has a similar function in rice, a transgenic homozygous rice line with the barley Hvsusiba2 was created previously. The cultivar was used in this study to compare its starch content and amylopectin synthase activity with the wild type. The results showed that the transgenic line exhibited a higher starch contents in the mature grains and stalks, but not in the leaves or roots, than that in the wild type.Starch accumulation dynamics in the process of grain development were similar between the transgene line and the wild counterpart in the initial 12 days after flowering, but markedly increased 15 days after flowering in the transgenic line. It suggested that Hvsusiba2 might not start affecting the starch synthesis until after mid and late grain filling stages in rice. Furthermore, the activities of amylopectin synthases, such as SBEIIb, SBEI, isoamylase and amylase, were enhanced in the developing grains of transgenic line, but the SBEIIa and pullulanase were not affected by Hvsusiba2. It appeared that the heterologous expression of Hvsusiba2 could increase the starch content, and likely, the activities of several amylopectin synthases as well, in rice similar to what it does in barley.
-
Key words:
- Hvsusiba2 /
- rice /
- total starch /
- amylopectin synthases
-
图 1 转基因水稻SU-7发育种子和茎鞘及其野生型水稻的蛋白粗提物的非变性凝胶
注：A为SU-7和非转基因对照日本晴发育9 d的种子和茎鞘的粗蛋白提取物在非变性胶中分离的条带，凝胶中含0.2% (W/V) 马铃薯直链淀粉，凝胶在30℃活化8 h后，用3%的碘液染色10 min后的图像；B为染色10 min后凝胶用ddH2O漂洗2次后的图像。
Figure 1. Native PAGE of amylopectin synthases in crude protein extracts of developing grains and stalks of wild type and transgenic rice SU-7
表 1 异源表达Hvsusiba2水稻成熟种子总淀粉含量
Table 1. Starch content in grains of Hvsusiba2 rice and wild type
下载: 导出CSV
表 2 异源表达Hvsusiba2水稻种子发育过程中总淀粉含量
Table 2. Starch content in developing grains of Hvsusiba2 rice and wild type
(单位/%) 水稻株系 花后3 d 花后5 d 花后9 d 花后12 d 花后15 d Nipp 1.4±0.42a 2.27±0.95a 9.74±1.76a 12.61±2.59a 16.22±0.97b SU-7 1.18±0.03a 2.84±2.03a 7.34±0.97a 12.28±0.29a 23.31±1.11a
下载: 导出CSV
表 3 异源表达Hvsusiba2水稻植株根、茎和叶总淀粉含量
Table 3. Starch content in roots, stalks and leaves of Hvsusiba2 rice and wild type
(单位/%) 组织 Nipp SU-7 根 0.097±0.015 a 0.093±0.006 a 茎 0.64±0.1480 b 1.36±0.1102 a 叶 0.31±0.0416 a 0.31±0.0819 a
下载: 导出CSV
-
[1] 高振宇, 黄大年, 钱前.植物支链淀粉生物合成研究进展[J].植物生理与分子生物学学报, 2004, 30(5):489-495. http://www.cnki.com.cn/Article/CJFDTOTAL-ZWSI200405001.htm [2] 谢祝捷, 姜东, 戴廷波, 等.植物的糖信号及其对碳氮代谢基因的调控[J].植物生理学通讯, 2002, 38(4):399-405. http://www.cnki.com.cn/Article/CJFDTOTAL-ZWSL200204036.htm [3] CHUANXIN SUN, SARA PALMQVIST, HELENA OLSSON, et al. A Novel WRKY Transcription Factor, SUSIBA2, Participates in Sugar Signaling in Barley by Binding to the Sugar-Responsive Elements of the iso1 Promoter[J]. The Plant Cell, 2003, 15:2076-2092. doi: 10.1105/tpc.014597 [4] CHUANXIN SUN, ANNA-STINA H GLUND, HELENA OLSSON, et al. Antisense oligodeoxynucleotide inhibition as a potent strategy in plant biology:identification of SUSIBA2 as a transcriptional activator in plant sugar signaling[J]. The Plant Journal, 2005, 44(1):128-138. doi: 10.1111/tpj.2005.44.issue-1 [5] 胡昌泉. 水稻胚乳发育相关基因的筛选及功能分析[D]. 福州: 福建农林大学, 2011: 62-64. [6] NAKAMURA Y, SAKURAI A, INABA Y, et al. The fine Structure of Amylopectin in Endosperm from Asian Cultivated Rice can be largely Classified into two Classes[J]. Starch/Stärke, 2002, 54(3-4):117-131. doi: 10.1002/1521-379X(200204)54:3/4<117::AID-STAR117>3.0.CO;2-2 [7] GE VANDEPUTTE, JA DELCOUR. From sucrose to starch granule to starch physical behaviour-a focus on rice starch[J]. Carbohydrate Polymers, 2004, 58(3):245-266. doi: 10.1016/j.carbpol.2004.06.003 [8] 苏军, 张武君, 陈在杰.一种快速的水稻不同组织总蛋白提取方法[J].福建农业学报, 2013, 28(9):884-887. http://www.fjnyxb.cn/CN/abstract/abstract2251.shtml [9] MARION M, BRADFORD A. Rapid and Sensitive Method for the Quantitation of Microgram Quantities of Protein Utilizing the Principle of Protein-Dye Binding[J]. Analytical Biochemistry, 1976, 72:248-254. doi: 10.1016/0003-2697(76)90527-3 [10] 朱海江, 程方民, 王丰, 等.两种穗型粳稻穗内粒间直链淀粉含量变异与粒位分布特征[J].中国水稻科学, 2004, 18 (4):321-325. http://kns.cnki.net/KCMS/detail/detail.aspx?dbcode=CJFQ&dbname=CJFD2004&filename=ZGSK200404006&v=MjA3MTk4ZVgxTHV4WVM3RGgxVDNxVHJXTTFGckNVUkwyZll1ZHNGeXZoVmIvSlB5cllaYkc0SHRYTXE0OUZZb1I= [11] JENNIFER VANDOOREN, NATHALIE GEURTS, ERIK MARTENS, et al. Zymography methods for visualizing hydrolytic enzymes[J]. Nature methods, 2013, 10(3):211-220. doi: 10.1038/nmeth.2371 -
点击查看大图
计量
- 文章访问数: 1508
- HTML全文浏览量: 221
- PDF下载量: 230
- 被引次数: 0
