[Screening and identification of CKI1 upstream transcription regulators in Arabidopsis].
[Screening and identification of CKI1 upstream transcription regulators in Arabidopsis].
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DOI：
10.16288/j.yczz.18-314
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发表时间：
2019-05-20
期刊：
影响因子：
--
通讯作者：
Yu, Xiao Lin
中科院分区：
文献类型：
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作者：
Liu, Zhen Ning;Yuan, Li;Yu, Xiao Lin
拟南芥CKI 1（cytokinin independent 1）是一种参与双组分系统的组胺酸激酶蛋白，可以通过下游的组胺酸磷酸转移蛋白激活双组分信号传导，在中央细胞命运决定和胚囊发育调节中发挥重要作用。然而，CKI 1上游转录调控因子的研究仍然有限。在本研究中，启动子活性的不同片段进行了调查，CKI 1上游转录调控筛选和鉴定的酵母单杂交技术。结果表明，位于内含子区的F5/R2片段在胚囊中具有启动子活性，与CKI 1全长启动子一致。然后利用F5/R2片段的3个串联重复序列构建诱饵表达载体，并收集拟南芥雌蕊进行cDNA文库构建。利用酵母单杂交技术共筛选出226个阳性克隆，去除低质量重复序列后，得到66个可读序列，其中8个蛋白质可作为DNA结合蛋白。这些结果为研究CKI 1在转录调控网络中的分子功能提供了重要线索。
Arabidopsis CKI1 (cytokinin independent 1) is a histidine kinase protein involved in the two-component system, which can activate two-component signaling via the downstream histidine phospho-transfer proteins, playing the essential roles in central cell fate determination and development regulation in embryo sacs. However, studies on CKI1 upstream transcription regulators are still limited. In the present study, promoter activities with varying fragments were investigated, and CKI1 upstream transcription regulators were screened and identified by the yeast-one hybrid technique. Results indicated F5/R2 fragments located in the intron region showed promoter activities in embryo sacs, which is consistent with CKI1 full-length promoters. Then three tandem repeats of F5/R2 fragments were used to construct the bait expression vector, and Arabidopsis pistils were collected for cDNA library construction. Totally, 226 positive clones were screened by the yeast-one hybrid technique, 66 readable sequences were retrieved after removing sequences with low quality and redundant repeats, among which eight proteins could act as DNA-binding proteins. These results provided some important clues to study the molecular function of CKI1 in the transcription regulation network.