[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.
Wang
D N
DN
College of Life Science and Technology, Shanghai Jiao Tong University, Shanghai 200030, China.
Li
M F
MF
Zhen
E L
EL
Zhang
W J
WJ
Wu
X F
XF
D054421:Chemokine CCL21; D019742:Chemokines, CC / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D020133:Reverse Transcriptase Polymerase Chain Reaction
C515478:CCL21 protein, human; D054421:Chemokine CCL21; D019742:Chemokines, CC