Xiong Kang-Hui, Liang Qin-Chuan, Xiong Hua, Zou Chang-Xu, Gao Guo-Dong, Zhao Zhen-Wei, Zhang Hua
Department of Anatomy, Fourth Military Medical University, 710032, Xi'an City, China.
Biotechnol Lett. 2005 Nov;27(21):1713-7. doi: 10.1007/s10529-005-2736-3.
A dicistronic expression vector was constructed for Chinese hamster ovary (CHO) cells that produce both selectable marker-DHFR (dihydrofolate reductase) gene and recombinant antibody cDNA from a single primary transcript via differential splicing. The vector was derived from a pDHL vector and contained the human constant region cDNA so that any human-mouse chimeric antibodies could be expressed. The expression vector produced stable CHO cell clones that secreted nearly double the amount of chimeric antibodies than produced by conventional expression approaches, where the DHFR gene and relevant cDNA are controlled by separate transcription cassettes. Clones with increased expression of interested genes can be efficiently generated by selection in medium containing a gradually increasing amount of methotrexate. The dicistronic expression system using incomplete splicing DHFR gene strategy thus provides a convenient, high-level, and rapid expression of chimeric antibodies.
构建了一种用于中国仓鼠卵巢（CHO）细胞的双顺反子表达载体，该载体通过差异剪接从单个初级转录本产生可选择标记——二氢叶酸还原酶（DHFR）基因和重组抗体cDNA。该载体源自pDHL载体，并包含人恒定区cDNA，以便能够表达任何人和小鼠嵌合抗体。与传统表达方法（其中DHFR基因和相关cDNA由单独的转录盒控制）相比，该表达载体产生的稳定CHO细胞克隆分泌的嵌合抗体量几乎是其两倍。通过在含有逐渐增加量甲氨蝶呤的培养基中进行选择，可以有效地产生感兴趣基因表达增加的克隆。因此，使用不完全剪接DHFR基因策略的双顺反子表达系统为嵌合抗体提供了一种方便、高水平且快速的表达方法。
