Shirai T, Shimizu N, Shiojiri S, Horiguchi S, Ito H
Life Science Research Laboratories, Asahi Chemical Industry Co., Ltd., Shizuoka, Japan.
DNA. 1988 Apr;7(3):193-201. doi: 10.1089/dna.1988.7.193.
The mouse tumor necrosis factor (TNF) gene was isolated from a mouse genomic library. The entire sequence of the gene was determined using both an automated DNA sequencer with improved primer extension reaction conditions as well as the standard radioisotopic method. Comparison of the nucleotide sequence of the gene with that of mouse TNF cDNA showed that the mouse gene consists of four exons, like rabbit and human TNF genes. There is strong nucleotide sequence homology in the 5'-flanking region among the mouse, rabbit, and human TNF genes, suggesting that the mechanisms regulating TNF gene expression are highly conserved. Direct expression of mature mouse TNF was achieved using a plasmid constructed by site-directed mutagenesis. Purified mouse TNF produced in Escherichia coli showed cytotoxicity to mouse L cells.
从小鼠基因组文库中分离出小鼠肿瘤坏死因子（TNF）基因。使用具有改进引物延伸反应条件的自动DNA测序仪以及标准放射性同位素方法确定了该基因的完整序列。将该基因的核苷酸序列与小鼠TNF cDNA的序列进行比较，结果表明，与兔和人TNF基因一样，小鼠基因由四个外显子组成。小鼠、兔和人TNF基因的5'侧翼区域存在很强的核苷酸序列同源性，这表明调节TNF基因表达的机制高度保守。通过定点诱变构建的质粒实现了成熟小鼠TNF的直接表达。在大肠杆菌中产生的纯化小鼠TNF对小鼠L细胞显示出细胞毒性。
