中文摘要：

目的建立研究DNA-蛋白质相互作用的新方法。方法选择6只SPF级BALB／c小鼠，模型组经尾静脉注射脂多糖（LPS）25mg／kg，使平均动脉压（MAP）降至40mmHg（1mmHg=0．133kPa）造成内毒素休克，然后迅速处死，正常对照组同期处死，取肝脏组织。利用生物素一链亲和素系统结合磁珠分离技术筛选内毒素诱导基因（LIG）启动子的结合蛋白。用聚合酶链反应（PCR）扩增末端带生物素标记的LIG启动子探针，提取动物肝脏组织胞核蛋白，与制备的LIG启动子探针进行孵育，再以标记有链亲和素的磁珠分离LIG启动子-蛋白反应复合物，使用不同浓度NaCl洗脱结合的蛋白，使用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS—PAGE）分离样品蛋白，并对凝胶进行银染显色，比较模型组与正常对照组的差异条带。结果两组胞核蛋白中共观察到15条差异条带。分析差异条带显示，与正常对照组比较，在低亲和力作用水平，LPS作用后有4个蛋白开始与LIG启动子作用力增强，而有1个蛋白则同启动子的相互作用减弱；在DNA-蛋白质高亲和力作用水平，则有9个蛋白因LPS刺激而同DNA的结合力增强，有1个蛋白同启动子的相互作用消失。结论成功建立了生物素-链亲和素结合磁珠分离技术的新方法，为研究DNA-蛋白质相互作用开拓了新思路，并从“转录调控组学”角度深入理解基因表达调控机制奠定了基础。

英文摘要：

Objective To establish a new method for the study of DNA-protein interaction. Methods Six SPF BALB/c mice were divided into two groups, the lipopolysaccharide （LPS）-treated group （n =3） and the normal control group （n=3）. The LPS-treated mice were subjected to tail vein-injection of LPS （25 mg/kg） to reproduce an endotoxic shock model. The biotin-labeled DNA probe for the LPS inducible gene （LIG） promoter was amplified by polymerase chain reaction （PCR）. The cell nuclear extracts from liver of mice were prepared and mixed together. DNA pull-down assays were performed by using magnetic beads conjugated with streptavidin. Proteins binding on the beads were eluted by salts with different concentrations. The samples were resolved by sodium dodecyl sulfate polyacrylamide gel electrophoresis （SDS-PAGE） and displayed by silver-stain to compare differential bands between two groups. Results Fifteen proteins in the nuclear extracts were found to be different between the mice of two groups. Analysis of the differential bands of SDS-PAGE displayed that, compared with normal control group, there were four proteins increased and one protein declined in the mice treated with LPS when eluted with low salt. While under the condition of high affinity elution, there were nine proteins increased and one disappeared after LPS treatment. Conclusion In this study, a novel method was set up by combining the biotin-streptavidin system and magnetic beads isolation technique, providing a new way to study DNA-protein interaction. This strategy is helpful for further understanding the regulation of gene expression with a view of ＂omics＂.