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

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213bp和417bp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、Lec2、Fus3、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Lec2在热激24h后上调表达,而常温培养下24h后没有变化;BcHSP、HSP18.2、Fus3和HSP17.6在热激24h后和常温下24h都上调表达;PAP14在热激24h后和常温下24h表达下调。

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

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 non-heat-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.