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构建白血病抑制因子(LIF)表达质粒pEDr-LIF并通过磷酸钙-DNA共沉淀的方法转入CHO-dhfr细胞。使用不同浓度的MTX筛选高表达株。获得的最高表达株经ELISA试剂盒检测上清中LIF含量3天内达到0.2μg/ml。Southern blot检测表明LIF cDNA已经稳固整合到了宿主细胞的基因组DNA中。MTT试验表明CHO-LIF的培养上清可以显著地抑制U937的细胞增殖。
The leukemia inhibitory factor (LIF) expression plasmid pEDr-LIF was constructed and transferred into CHO-dhfr cells by calcium phosphate-DNA co-precipitation. Highly expressed strains were screened using different concentrations of MTX. The highest expression strain obtained was detected by an ELISA kit to determine that the LIF content in the supernatant reached 0.2 μg/ml within 3 days. Southern blot analysis showed that the LIF cDNA has been stably integrated into the genomic DNA of the host cell. The MTT assay showed that the culture supernatant of CHO-LIF could significantly inhibit U937 cell proliferation.