[The study of BL21(DE3)/PET-11 as an expression system of human IL-2].
The coding region of hIL-2 cDNA is specifically amplified by PCR. A Nde I site is introduced into its 5'-end and its 3'-non coding region is partly deleted. After filling-in by Klenow enzyme, the DNA fragment is cloned into Sma I site of PUC18. Eight white colonies are screened and one (C1) contains Nde I site. DNA sequencing shows that non-specific mutation has not been found in its coding region. The DNA fragment digested by BamH I, Nde I of C1 plasmid is recovered and subcloned into PET-11 and transfered into BL21 (DE3). After IPTG induction, the Bacteria lysate is run directly on SDS-PAGE. Western blot test shows a specific rhIL-2 McAb binding band. The molecular weight is about 17000. Laser scanning indicates that the absorption area of IL-2 is about 5% of the total area.
Gong
M
M
Institute of Basic Medical Sciences, CAMS, Beijing.
Zhang
J
J
Liu
D
D
Zhang
S
S
Liu
P
P
D001483:Base Sequence; D018076:DNA, Complementary / Q000235:genetics; D004274:DNA, Recombinant / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D006801:Humans; D007376:Interleukin-2 / Q000096:biosynthesis* / Q000235:genetics; D008969:Molecular Sequence Data; D017422:Sequence Analysis, DNA
D018076:DNA, Complementary; D004274:DNA, Recombinant; D007376:Interleukin-2