[1]徐金龙,梁爽,郁飞.拟南芥细胞周期基因AtCDC5的功能研究及抗体制备[J].江苏农业学报,2019,(01):26-32.[doi:doi:10.3969/j.issn.1000-4440.2019.01.004]
　XU Jin-long,LIANG Shuang,YU Fei.Functional research and antibody preparation of Arabidopsis thaliana cell cycle gene AtCDC5[J].,2019,(01):26-32.[doi:doi:10.3969/j.issn.1000-4440.2019.01.004]
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拟南芥细胞周期基因AtCDC5的功能研究及抗体制备()
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江苏农业学报[ISSN:1006-6977/CN:61-1281/TN]

卷:
期数:
2019年01期
页码:
26-32
栏目:
遗传育种·生理生化
出版日期:
2019-02-26

文章信息/Info

Title:
Functional research and antibody preparation of Arabidopsis thaliana cell cycle gene AtCDC5
作者:
徐金龙; 梁爽; 郁飞
（西北农林科技大学生命科学学院，陕西杨陵712100）
Author(s):
XU Jin-long; LIANG Shuang; YU Fei
（College of Life Sciences, Northwest A&F University, Yangling 712100, China）
关键词:
细胞周期基因CDC5; 亚细胞定位; 原核表达; 抗体制备
Keywords:
cell division cycle 5; subcellular localization; prokaryotic expression; antibody preparation
分类号:
Q786
DOI:
doi:10.3969/j.issn.1000-4440.2019.01.004
文献标志码:
A
摘要:
MYB相关CDC5 (Cell division cycle 5)蛋白对于细胞周期G2期的正常进行是必须的，但是在植物中对这种蛋白质的研究还较少涉及。本试验测量比较野生型拟南芥（Col-0生态型）和AtCDC5 T-DNA插入缺失突变体（GK_278B09）的主根生长速度和长度，利用瞬时表达AtCDC5-GFP研究AtCDC5在拟南芥叶片叶肉细胞中的定位，同时利用AtCDC5 N端144个氨基酸在大肠杆菌中进行原核表达和纯化，免疫家兔获得针对AtCDC5的多克隆特异性抗体，从表型、细胞和蛋白质水平研究AtCDC5基因的功能。结果表明，GK_278B09的主根因发育受到抑制而生长速度缓慢；AtCDC5-GFP融合蛋白主要定位于细胞核中；经过分离得到的AtCDC5蛋白抗血清能够有效地检测出10 ng原核表达的AtCDC51-144抗原，并在原生质体AtCDC5过表达体系中检测出1条分子量约为120 000的条带，此条带与AtCDC5-GFP大小相近。本研究成功制备出AtCDC5蛋白的多克隆抗体，为进一步研究AtCDC5蛋白影响细胞周期从而影响植物生长发育提供了研究基础。
Abstract:
MYB related protein cell division cycle 5 (CDC5）is essential for the G2 phase of cell cycle, but the research about CDC5 in plant is few. In the present study, the main root length and growth rate of wild-type Arabidopsis thaliana (Col-0 ecotype) and AtCDC5 T-DNA insertion deletion mutant (GK_278B09) were compared, and the subcellular localization of the AtCDC5 protein was studied by transient expressed AtCDC5-GFP. Moreover, prokaryotic expression and were applied using 144 amino acids at the N-terminus of AtCDC5 in Escherichia coli. The polyclonal antibody against AtCDC5 was obtained by immunizing rabbits. The function of AtCDC5 was researched at levels of phenotype, cell and protein. The results showed that the main root development of GK_278B09 was inhibited so that the rate of growth was lower than Col. The AtCDC5-GFP fusion protein was located in the nuclei. The separated serum of AtCDC5 could test 10 ng of AtCDC5 antigen, and a band with themolecular weight of 120 000 was detected in the protoplast system. Furthermore, the size of the band was similar to that of AtCDC5-GFP. The polyclonal antibody against AtCDC5 provides resource for the research on the function of AtCDC5 in plant growth.

参考文献/References:

[1]邹向阳，李连宏.细胞周期调控与肿瘤[J].国际遗传学杂志, 2006（29）：70-73.
[2]LIN Z Q , YIN K Q, ZHU D L, et al. AtCDC5 regulates the G2 to M transition of the cell cycle and is critical for the function of Arabidopsis shoot apical meristem[J]. Cell Research, 2007, 17(9)：815-828.
[3]CHEN Y H, YANG X Y, HE K, et al. The MYB transcription factor superfamily of Arabidopsis：expression analysis and phylogenetic comparison with the rice MYB family[J].Plant Molecular Biology, 2006, 60：107-124.
[4]HIRAYAMA T, SHINOZAKI K. A cdc5+ homolog of a higher plant, Arabidopsis thaliana[J].PNAS, 1996, 93(23)：13371-13376.
[5]LIN Z Q, YIN K Q, WANG X X, et al. Virus induced gene silencing of AtCDC5 results in accelerated cell death in Arabidopsis leaves[J].Plant Physiol Biochem, 2007, 45(1)：87-94.
[6]MONAGHAN J, XU F, XU S H, et al. Two putative RNA-binding proteins function with unequal genetic redundancy in the MOS4-associated complex[J].Plant Physiol, 2010, 154(4)：1783-1793.
[7]PALMA K, ZHAO Q G, CHENG Y T, et al. Regulation of plant innate immunity by three proteins in a complex conserved across the plant and animal kingdoms[J].Genes Dev, 2007, 21(12)：1484-1493.
[8]YOO S D, CHO Y H, SHEEN J. Arabidopsis mesophyll protoplasts：a versatile cell system for transient gene expression analysis[J].Nat Protoc, 2007, 2(7)：1565-1572.
[9]余璐璐,龚绒雪,吕林涛，等. 拟南芥氰丙氨酸合酶CYS-C1基因扩增及多克隆抗体制备[J].江苏农业学报，2017, 33(6)：1235-1241.
[10]杨雄,刘秀侠,南昊，等. HIV-1可溶性单链抗体b12-scFv的表达与活性分析[J].西北农林科技大学学报, 2014, 42(5)：205-210.

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备注/Memo

备注/Memo:
收稿日期：2018-03-08 基金项目：国家自然科学基金项目（31741010） 作者简介：徐金龙（1992-），男，山东德州人，硕士研究生，主要从事拟南芥突变体的筛选研究。（E-mail）xujinlongxy@163.com 通讯作者：郁飞，（E-mail）flyfeiyu@gmail.com
更新日期/Last Update: 2019-02-27