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目的:应用酵母双杂交系统从人外周血白细胞cD-NA文库中筛选与转录因子Foxp3相互作用的蛋白,为进一步研究Foxp3的转录调控机制奠定基础。方法:首先构建pG-BKT7-Foxp3△2酵母双杂交诱饵载体,转化AH109酵母细胞,检测其毒性及自激活作用;然后将诱饵质粒与人外周血白细胞cDNA文库共转AH109酵母细胞,筛选了与Foxp3△2存在相互作用的蛋白。结果:成功构建了pGBKT7-Foxp3△2酵母表达载体,经转染AH109酵母细胞,无有毒性,无自激活作用。获得了40个阳性克隆,经生物信息学分析,其中9个具有开放读框。结论:应用酵母双杂交技术筛选出一组可与Foxp3△2相互作用的候选蛋白,为进一步研究Foxp3△2功能奠定了基础。
OBJECTIVE: To screen the protein interacting with transcription factor Foxp3 from cD-NA library of human peripheral blood leukocytes by yeast two-hybrid system and lay a foundation for further study on the transcriptional regulation of Foxp3. Methods: The pG-BKT7-Foxp3 △ 2 yeast two-hybrid bait vector was constructed and transformed into AH109 yeast cells to detect its toxicity and self-activation. Then the bait plasmid and human peripheral blood leukocyte cDNA library were co-transfected into AH109 yeast cells, Foxp3 △ 2 There are interacting proteins. Results: The pGBKT7-Foxp3Δ2 yeast expression vector was successfully constructed and transfected into AH109 yeast cells without toxicity and self-activation. Forty positive clones were obtained, of which 9 were open reading frames by bioinformatics analysis. Conclusion: A set of candidate proteins that can interact with Foxp3 △ 2 was screened by yeast two-hybrid technique, which laid the foundation for further study on Foxp3 △ 2 function.