一个水稻小G蛋白OsRan1的克隆及表达分析

摘要/Abstract

摘要： 逆境对于水稻产量和品质有很大影响，研究在逆境下水稻基因的表达变化对于研究水稻抗逆有着重要意义。该实验运用荧光差异显示(Fluorescence Differential Display, FDD)技术，从低温处理的水稻中克隆到OsRAN1基因并通过半定量PT-PCR分析该基因在低温和高盐逆境下的表达。该基因cDNA序列全长为984bp，含有一个编码221个氨基酸残基的开放阅读框，推测分子量为25.0 KD，此氨基酸序列具有GTPase活性区域，属于小G蛋白Ran亚家族，通过氨基酸相似性比对发现，Ran家族氨基酸序列高度保守，OsRAN1与TaRAN1和AtRAN1相似性分别达到98.19%和92.76%。通过半定量PT-PCR分析，发现在低温胁迫下，OsRAN1在根和叶鞘中表达量均有增加，尤其是高盐胁迫下根中OsRAN1表达量有迅速升高然后降低的变化。推测该基因在水稻逆境应答中起着重要作用。

关键词: 低山丘陵区, 低山丘陵区, 基本农田, 土地利用总体规划, 广安

Abstract: The disadvantage circumstance is an important factor in impacting on the yield and quality of rice .The expression analysis of gene under stress helps a lot in studying of stress resistance of rice. A rice gene, OsRAN1, was isolated from rice treated with cold-stress by the fluorescence differential display (FDD) screening method，and its expression pattern was analyzed by simi-quantitative RT-PCR analysis under cold and salt stress. Its cDNA sequence (984bp) contains an ORF encoding a 221 amino acids protein (25.0KD)，and the protein belongs to Ran subfamily of the small G protein, for containing the typical GTPase structure. At the amino-acid sequence level, OsRAN1 exhibit high homology to TaRAN1 and AtRAN1 (98.19% and 92.76%). By simi-quantitative RT-PCR analysis, the expression of OsRAN1 at the transcription level was increased under the low temperature. Especially under high salt condition, the expression level in root shows a process of increase, and then decrease. We suggest the OsRAN1 gene could play an important role of stress response in rice.

席江,杜志如,徐永菊,张琦,万佳,江斌,徐正君,. 一个水稻小G蛋白OsRan1的克隆及表达分析[J]. 中国农学通报, 2008, 24(4): 109-114.

Xi Jiang, Du Zhiru, Xu Yongju , Zhang Qi , Wan Jia, Jiang Bin, Xu Zhengjun,. Isolation and Characterization of a Rice Gene OsRan1 Coding a Small G Protein[J]. Chinese Agricultural Science Bulletin, 2008, 24(4): 109-114.

参考文献

[1] Drivas, G..T., A. Shih, E. Coutavas, et al. Characterization of four novel ras-like genes expressed in a human teratocarcinoma cell line. Mol Cell Biol., 1990, 10(4): 1793-1798.
[2] Coutavas, E., C.-M. Hsieh, G. Drivas, et al. Tissue-specific expression of Ran isoforms in the mouse. Mamm Genome, 1994, 5(10): 623-628.
[3] Marshall, M.S. . The effector interactions of p21ras. Trends Biochem. Sci, 1993, 18: 250-254.
[4] Sazer, S , M. Dasso. The ran decathlon: multiple roles of Ran. J Cell Sci., 2000, 113 (7): 1111-1118.
[5] Yasuda, Y., Y. Miyamoto, T. Saiwaki, et al. Mechanism of the stress-induced collapse of the Ran distribution. Exp Cell Res., 2006, 312(4): 51220.
[6] Go¨ rlich D, Kutay U .Transport between the cell nucleus and the cytoplasm. Annu Rev Cell Dev Biol , 1999, 15: 607-660.
[7] Nachury, M. V., K. Weis. The direction of transport through the nuclear pore can be inverted. Proc. Natl. Acad. Sci. , 1999, 96: 9622-9627
[8] Hetzer M, Gruss O J, Mattaj I W. The Ran GTPase as a marker of chromosome position in spindle formation andnuclear envelope assembly. Nat Cell Biol, 2002, 4: 177-4184.
[9] Zhang, C. , P.R. Clarke. Chromatin-independent nuclear envelope assembly induced by Ran GTPase in Xenopus egg extracts. Sci., 2000, 288(5470):1429-1432.
[10] Ach, R.A. , Gruissem, W. A small nuclear GTP-binding protein from tomato suppresses a Schizosaccharomycespombe cell-cycle mutant. Proc. Natl Acad. Sci., 1994, 91: 5863-5867.
[11] Haizel T, Merkle T, Pay A, et al. Characterization of proteins that interact with the GTP-bound form of the regulatory GTPase Ran in Arabidopsis. Plant J. , 1997, 11: 93-103.
[12] Kim S H, Arnold D, Lloyd A,et al. Antisense expression of an Arabidopsis Ran binding protein renders transgenic roots hypersensitive to auxin and alters auxin-induced root growth and development by arresting mitotic progress. Plant Cell, 2001, 13: 2619-2630.
[13] Xin W, Yunyuan X., Ye, H. , et al. Overexpression of RAN1 in Rice and Arabidopsis Alters Primordial Meristem, Mitotic Progress, and Sensitivity to Auxin1, Plant Physiol., 2006, 140:91-101.
[14] Wladimir I L T, David C. B. Physical Association of the NB-LRR Resistance Protein Rx with a Ran GTPase–Activating Protein Is Required for Extreme Resistance to Potato virus X. The plant cell, 2007, 19:1682-1694.
[15] Logemann, J., J. Schell, L. Willmitzer. Improved method for the isolation of RNA from plant tissues. Anal Biochem, 1987, 163(1): 16-20.
[16] Stephanie A R, Karen M L, Ian G M.. The C Terminus of the Nuclear RAN/T4 GTPase Stabilizes the GDP-bound State and Mediates Interaction with RCC1, RAN-GAP, and HTF9A/RANBP1. Proc.Natl.Acad.Sci. , 1995, 270:14405-14411.
[17] F.R. Bischoff, H. Ponstingl. Catalysis of guanine nucleotide exchange on Ran by the mitotic regulator RCC1. Nature , 1991, 354 : 80-82.
[18] F.R. Bischoff, H. Krebber, T. Kempf, et al. Human RanGTPase-activating protein RanGAP1 is a homologue of yeast Rna1p involved in mRNA processing and transport. Proc. Natl. Acad. Sci. 1995, 92: 1749-1753.
[19] Y. Miyamoto, T. Saiwaki, J. Yamashita, et al. Cellular stresses induce the nuclear accumulation of importin a and cause a conventional nuclear import block. J. Cell Biol., 2004, 165: 617- 623.
[20] Susanne B, Karin S, Karl-Josef D. Characterization of a small GTP-binding protein of the rab 5 family in Mesembryanthemum crystallinum with increased level of expression during early salt stress. Plant.Mol.Bio, 2000, 42: 923-936.