《植物生理学报》 2009, 45(12): 1191-1196

大白菜中小孢子胚胎发生相关基因cDNA 片段的克隆与表达分析

汪维红^1,2, 于拴仓², 许明¹, 赵岫云², 张凤兰^2,*, 余阳俊², 张德双²

1 沈阳农业大学园艺学院, 沈阳110866; 2 北京市农林科学院蔬菜研究中心, 北京100097

通信作者：张凤兰；E-mail: zhangfenglan@nercv.org；Tel: 010-51503038

摘　要：

以易出胚大白菜‘09C3’ 为材料, 克隆到7 个小孢子胚胎发生相关候选基因的cDNA核心片段, 并采用半定量RT-PCR方法研究了这些基因在游离小孢子热激处理和培养期间的表达。结果表明, 获得的 7 个基因的核心片段长度在 213 bp 和417 bp 之间, 克隆号分别为 C31、C33、99H6、99H8、EV14、EX01 和 C35。NCBI 进行 Blastn 和 Blastx 分析结果显示, 这些片段与源基因 HSP70、BcHSP、Lec 2、Fus 3、PAP14、HSP17.6 和 HSP18.2 的同源性均超过 80%。半定量 RT-PCR 结果表明, HSP70 和 Lec 2 在热激 24 h 后上调表达, 而常温培养下 24 h 后没有变化; BcHSP、HSP18.2、Fus 3 和 HSP17.6 在热 激 24 h 后和常温下 24 h 都上调表达; PAP14 在热激 24 h 后和常温下 24 h 表达下调。

关键词：大白菜; 小孢子胚胎发生; 热激; 半定量RT-PCR

收稿：2009-10-26   修定：2009-11-17

资助：国家农业行业计划(nyhyzx07-007)、北京市科技计划项目(Z09090501040902)和现代农业产业技术体系建设专项资金。

Cloning and Expression Analysis of cDNA Fragments of Genes Related to Microspore Embryogenesis in Chinese Cabbage

WANG Wei-Hong^1,2, YU Shuan-Cang², XU Ming¹, ZHAO Xiu-Yun², ZHANG Feng-Lan^2,*, YU Yang-Jun², ZHANG De-Shuang²

¹College of Horticulture, Shenyang Agricultural University, Shenyang 110866, China; ²Beijing Academy of Agriculture and Forestry Science (BAAFS), Beijing 100097, China

Corresponding author: ZHANG Feng-Lan; E-mail: zhangfenglan@nercv.org; Tel: 010-51503038

Abstract:

The highly embryogenic inbred line ‘09C3’ was used to clone cDNA fragments of seven candidate genes involving in microspore embryogenesis of Chinese cabbage, and the expression of these genes were studied by semi-quantitative RT-PCR to monitor the effect of 24 h heat-shock and gene behavior after 24 h nonheat-shock culture. The length of seven obtained fragments ranged from 213 bp to 417 bp (the cloning IDs were C31, C33, 99H6, 99H8, EV14, EX01 and C35, respectively). By Blastn and Blastx on the NCBI, these fragments showed over 80% in amino acid identity against HSP70, BcHSP, Lec 2, Fus 3, PAP14, HSP17.6 and HSP18.2, respectively. The results from semi-quantitative RT-PCR showed that HSP70 and Lec 2 were upregulated after 24 h heat shock, while no changes were found in normal culture. BcHSP, HSP18.2, Fus 3 and HSP17.6 were up-regulated under both treatments, while PAP14 was down-regulated.

Key words: Chinese cabbage; microspore embryogenesis; heat shock; semi-quantitative RT-PCR