次组淋巴趋势化因子SLC的克隆及其在原核系统中的表达

以中国人的淋巴细胞组织为材料提总RNA，用RT－PCR的方法扩增了次级淋巴组织趋化因子（Secondary lymphoid-tissue chemokine,SLC)的成熟肽基因，序列分析表明，我们克隆的SLC基因与文献报道的仅有一个核苷酸的差异,且并不影响氨基酸的编码。将PLC的cDNA插入含T7启动子的表达载体pET-28a( ）中构建重组质粒pET-SLC,转化大肠杆菌BL21（DE3）筛选表达菌株，表达菌株经1mmol/LIPTG诱导表达3－5h后，超 声破菌，离心后将上清进行SDS－PAGE，可以看到在18KD左右处有明显的表达条带，用Ni^2 亲和层析柱纯化表达物，纯度表达90％以上。

Cloning of Secondary Lymphoid-tissue Chemokine (SLC) and Its Expression in Prokaryotic System

The total RNA from lymphoid tissue in Chinese was extracted, and the gene encoding the mature peptide of secondary lymphoid-tissue chemokine (SLC) was cloned by RT-PCR. Nucleotide sequence analysis showed that there is only one nucleotide different from that reported, but it doesn't alter the amino acid encoded. The SLC cDNA was inserted into an expression vector pET-28a(+) under T7 promoter and constructed recombinant plasmid pET28a-SLC. pET28a-SLC was transformed to E. coli BL21(DE3) and the expression strain was gotten. After inducing with IPTG for 3-5 hours the bacterium were sonicated. After centrifuging the supernatant was analysed by SDS-PAGE. An obvious expression band about 18 kD can be seen. The expressed product was purified by Ni2+ affinity chromatography column, and the purity is up to 90 percent.