|本期目录/Table of Contents|

[1]杜京尧,尚飞,王高华,等.OsRhoGDI2过表达转基因水稻的筛选鉴定及外源基因拷贝数的初步分析[J].江苏农业科学,2019,47(14):50-54.
　Du Jingyao,et al.Screening and identification of OsRhoGDI2 overexpressed transgenic rice and preliminary analysis of copy number of foreign gene[J].Jiangsu Agricultural Sciences,2019,47(14):50-54.
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OsRhoGDI2过表达转基因水稻的筛选鉴定
及外源基因拷贝数的初步分析(PDF)
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《江苏农业科学》[ISSN:1002-1302/CN:32-1214/S]

卷:
第47卷
期数:
2019年第14期
页码:
50-54
栏目:
生物技术
出版日期:
2019-08-10

文章信息/Info

Title:
Screening and identification of OsRhoGDI2 overexpressed transgenic rice and preliminary analysis of copy number of foreign gene
作者:
杜京尧;  尚飞;  王高华;  梁卫红
河南师范大学生命科学学院，河南新乡 453007
Author(s):
Du Jingyao; et al
关键词:
OsRhoGDI2; 转基因水稻; 表达分析; 拷贝数; 外源基因
Keywords:
-
分类号:
S511.03
DOI:
-
文献标志码:
A
摘要:
水稻Rho GDP解离抑制基因OsRhoGDI2是从幼穗中分离出的功能未知基因。为鉴定该基因的功能，笔者所在实验室前期构建了植物过表达载体pCAMBIA1302-OsRhoGDI2-GFP，并对水稻进行了遗传转化。对OsRhoGDI2过表达转基因水稻T2代进行筛选和鉴定，采用PCR技术鉴定转基因植株，采用半定量RT-PCR和实时荧光定量PCR检测OsRhoGDI2在转基因水稻中的表达水平，结果显示，其中6个株系为过表达转基因植株，OsRhoGDI2表达水平上调1.69～13.35倍。为检测外源基因在转基因水稻中的拷贝数，分别以蔗糖磷酸合成酶基因SPS和潮霉素抗性基因HYG为内参基因和标记基因，采用实时荧光定量PCR(qPCR)技术结合内参基因和标记基因的标准曲线进行分析，结果显示在所检测的6个转基因株系中，外源基因的拷贝数均为1，提示已经获得稳定遗传的OsRhoGDI2过表达转基因水稻，为后续OsRhoGDI2基因的功能研究奠定基础。
Abstract:
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参考文献/References:

［1］Kost B，Lemichez E，Spielhofer P，et al. Rac homologues and compartmentalized phosphatidylinositol 4，5-bisphosphate act in a common pathway to regulate polar pollen tube growth［J］. Journal of Cell Biology，1999，145(2)：317-330.
［2］Molendijk A J，Bischoff F，Rajendrakumar C S，et al. Arabidopsis thaliana Rop GTPases are localized to tips of root hairs and control polar growth［J］. EMBO Journal，2001，20(11)：2779-2788.
［3］Jones M A，Shen J J，Fu Y，et al. The Arabidopsis Rop2 GTPase is a positive regulator of both root hair initiation and tip growth［J］. The Plant Cell，2002，14(4)：763-776.
［4］Fu Y，Wu G，Yang Z B. Rop GTPase-dependent dynamics of tip-localized F-actin controls tip growth in pollen tubes［J］. Journal of Cell Biology，2001，152(5)：1019-1032.
［5］Fu Y，Li H，Yang Z B. The ROP2 GTPase controls the formation of cortical fine F-actin and the early phase of directional cell expansion during Arabidopsis organogenesis［J］. The Plant Cell，2002，14(4)：777-794.
［6］Delmer D P，Pear J R，Andrawis A，et al. Genes encoding small GTP-binding proteins analogous to mammalian rac are preferentially expressed in developing cotton fibers［J］. Molecular & General Genetics，1995，248(1)：43-51.
［7］Trotochaud A E，Hao T，Wu G，et al. The CLAVATA1 receptor-like kinase requires CLAVATA3 for its assembly into a signaling complex that includes KAPP and a Rho-related protein［J］. The Plant Cell，1999，11(3)：393-406.
［8］Kawasaki T，Henmi K，Ono E，et al. The small GTP-binding protein rac is a regulator of cell death in plants［J］. Proceedings of the National Academy of Sciences of the United States of America，1999，96(19)：10922-10926.
［9］Kawasaki T，Koita H，Nakatsubo T，et al. Cinnamoyl-CoA reductase，a key enzyme in lignin biosynthesis，is an effector of small GTPase Rac in defense signaling in rice［J］. Proceedings of the National Academy of Sciences of the United States of America，2006，103(1)：230-235.
［10］Fujiwara M，Umemura K，Kawasaki T，et al. Proteomics of Rac GTPase signaling reveals its predominant role in elicitor-induced defense response of cultured rice cells［J］. Plant Physiology，2006，140(2)：734-745.
［11］Schultheiss H，Hensel G，Imani J，et al. Ectopic expression of constitutively activated RACB in barley enhances susceptibility to powdery mildew and abiotic stress［J］. Plant Physiology，2005，139(1)：353-362.
［12］Dermardirossian C，Bokoch G M. GDIs：central regulatory molecules in Rho GTPase activation［J］. Trends in Cell Biology，2005，15(7)：356-363.
［13］Dovas A，Couchman J R. RhoGDI：multiple functions in the regulation of Rho family GTPase activities［J］. The Biochemical Journal，2005，390：1-9.
［14］Klahre U，Becker C，Schmitt A C，et al. Nt-RhoGDI2 regulates Rac/Rop signaling and polar cell growth in tobacco pollen tubes［J］. Plant Journal，2006，46(6)：1018-1031.
［15］Bischoff F，Vahlkamp L，Molendijk A，et al. Localization of AtROP4 and AtROP6 and interaction with the guanine nucleotide dissociation inhibitor AtRhoGDI1 from Arabidopsis［J］. Plant Molecular Biology，2000，42(3)：515-530.
［16］Kieffer F，Elmayan T，Rubier S，et al. Cloning of Rac and Rho-GDI from tobacco using an heterologous two-hybrid screen［J］. Biochimie，2000，82(12)：1099-1105.
［17］梁卫红，唐朝荣，吴乃虎. 两种水稻GDP解离抑制蛋白基因的分离及特征分析［J］. 中国生物化学与分子生物学报，2004，20(6)：785-791.
［18］Carol R J，Takeda S，Linstead P，et al. A RhoGDP dissociation inhibitor spatially regulates growth in root hair cells［J］. Nature，2005，438(770)：1013-1016.
［19］Wu Y X，Zhao S J，Tian H，et al. CPK3-phosphorylated RhoGDI1 is essential in the development of Arabidopsis seedlings and leaf epidermal cells［J］. Journal of Experimental Botany，2013，64(11)：3327-3338.
［20］彭威风，梁卫红. 水稻OsRhoGDI2蛋白生物信息学分析及亚细胞定位研究［J］. 生物技术通报，2010(5)：82-86，92.
［21］Huang J J，Zhang J，Hao Y F，et al. Distinct expression patterns of the GDP dissociation inhibitor protein gene (OsRhoGDI2) from Oryza sativa during development and abiotic stresses［J］. Biologia，2016，71(11)：1230-1239.
［22］Livak K J，Schmittgen T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCT method［J］. Methods，2001，25(4)：402-408.
［23］杨立桃，赵志辉，丁嘉羽，等. 利用实时荧光定量PCR方法分析转基因水稻外源基因拷贝数［J］. 中国食品卫生杂志，2005，17(2)：140-144.
［24］Ding J Y，Jia J W，Yang L T，et al. Validation of a rice specific gene，sucrose phosphate synthase，used as the endogenous reference gene for qualitative and real-time quantitative PCR detection of transgenes［J］. Journal of Agricultural&Food Chemistry，2004，52(11)：3372-3377.
［25］Flavell R B. Inactivation of gene expression in plants as a consequence of specific sequence duplication［J］. Proceedings of the National Academy of Sciences of the United States of America，1994，91(9)：3490-3496.
［26］Vaucheret H，Béclin C，Elmayan T，et al. Transgene-induced gene silencing in plants［J］. The Plant Journal，1998，16(6)：651-659.

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备注/Memo

备注/Memo:
收稿日期：2018-03-13
基金项目：国家自然科学基金(编号：31171182、U1704101)；河南省高校科技创新团队支持计划(编号：15IRTSTHN020)。
作者简介：杜京尧(1990—)，男，河南禹州人，硕士研究生，主要从事植物发育分子生物学研究。E-mail：dujingyao1990@126.com。
通信作者：梁卫红，博士，教授，主要从事植物发育分子生物学研究。Tel：(0373)3326340；E-mail：liangwh@htu.cn。
更新日期/Last Update: 2019-07-20