Abstract: To further explore the mechanism of ZjNAC2 in stress response,this study screened proteins interacting with ZjNAC2 transcription factors in Zoysia japonica by yeast two-hybrid technique. The gene sequence of Z. japonica ZjNAC2 was obtained by cloning and successfully constructed into the bait vector pGBKT7-ZjNAC2A High-quality full-length cDNA library of Z. japonica was also constructed. In the yeast two-hybrid screening experiment,we found 30 positive colonies and identified 20 proteins that interacted with ZjNAC2 by sequencing. The results of NR annotation analysis showed that 20 annotated genes were aligned to 9 homologous species,and the annotated genes annotated in Sorghum bicolor were the most. GO annotation results indicated that these genes can be classified in three major categories:cellular components,molecular functions,and biochemical processes. Four candidate interaction proteins related to disease resistance and plant growth and development were initially screened and selected,which laid a foundation for further research on the downstream regulation mechanism of ZjNAC2 in Z. japonica.

Key words: ZjNAC2, Transcriptional autoactivation, Yeast two-hybrid, Candidate gene

摘要： 为了进一步探究ZjNAC2在胁迫响应中的作用机制，本试验运用酵母双杂技术筛选与日本结缕草（Zoysia japonica）中的ZjNAC2转录因子相互作用的蛋白。通过克隆获得了日本结缕草ZjNAC2的基因序列并成功构建了诱饵载体pGBKT7-ZjNAC2，同时构建高质量的日本结缕草的均一化酵母杂交全长文库。通过酵母双杂筛库的实验筛选到30个阳性菌落，其测序表明获得了20个与ZjNAC2互作的蛋白。NR（Non-redundant protein sequences）注释分析结果显示，20个注释基因NR同源物种比对到9个物种，其中比对到高粱（Sorghum bicolor）中的注释基因最多。GO（Gene Ontology）注释结果表明这些基因可分为细胞组分、分子功能和生化过程3大类。本研究初步筛选到4个与抗病、植物生长发育过程相关的候选互作蛋白，为进一步探究ZjNAC2在日本结缕草中的下游调控机制奠定了基础。

关键词: ZjNAC2, 转录自激活, 酵母双杂交, 候选基因