全长与成熟形式IL-37b的真核表达载体的构建与研究

Construction and characterization of a eukaryotic expression vector of full-length and mature IL-37b

目的构建全长与成熟形式IL-37b的真核表达载体p EGFP N1/IL-37b,检测二者在RAW 264.7细胞中的表达情况。方法以含IL-37b全长编码区基因的质粒p UBC/IL-37b为模板,构建能够表达全长与成熟形式IL-37b的真核表达载体。将构建的重组质粒p EGFP N1/IL-37b转染到RAW 264.7细胞中,通过western blot和共聚焦显微镜检测全长与成熟形式的IL-37b的表达情况,并通过real-time PCR检测全长与成熟形式IL-37b对LPS诱导的IL-6表达的抑制作用。结果构建的p EGFP N1/IL-37b转染后能够在细胞中表达全长与成熟形式的IL-37b,并且能够抑制LPS诱导的IL-6的表达。结论成功构建了全长与成熟形式IL-37b的真核表达载体,为进一步研究IL-37b的炎症抑制作用与机制打好基础。

Objective To construct a eukaryotic expression vector p EGFP N1/IL-37 b of full-length and mature IL-37 b,and to detect the expression of both full-length and mature IL-37 b in RAW 264. 7 cells,a mouse macrophage cell line. Methods To construct the eukaryotic vectors of full-length and mature IL-37 b by using plasmid p UBC/IL-37 b as a template containing the coding region of IL-37 b full-length gene. To detect the expression of IL-37 b by western blot and confocal microscopy after transfected the recombinant plasmid into RAW 264. 7 cells,and to detect the inhibition of fulllength and mature IL-37 b on IL-6 production by real-time PCR. Results Eukaryotic vectors p EGFP N1/IL-37 b expressed full-length and mature IL-37 b after transfection in cells,which inhibited LPS-induced IL-6 production. Conclusions Eukaryotic vectors of full-length and mature IL-37 b can be successfully constructed,and lays a foundation for further study of anti-inflammation functions and mechanisms of IL-37 b.