Fig. 1
Generation of a conditional reversible KO mouse for IL-6. a Outline of the targeting strategy. Left: 5614 bp HpaI-PacI (pink; upstream) and a 5241 bp XhoI-NotI (orange; downstream) flanking Il6DIOex2 fragments. Right: the final product that consists of a PacI-SalI fragment with inverted exon 2 flanked by double lox sites, and three fragments containing Pgk-DTa and HSV-TK genes for negative selection and frt-flanked SvNeo gene for positive selection. b Southern blot for Il6 gene of the positive ES clones using the probe (left) revealed a ~11 kb band on the wild type allele and a ~8 kb band on the targeted allele, as expected (right); those clones with two bands were selected. c Left: The Il6 gene-modified sequence from IL6-DIO-KO mouse was sequenced by the Sanger method. The resulting sequence had 1361 bp and exon 1 (yellow), exon 2 (blue), lox2272 cassettes (green), loxP cassettes (purple), and FRT (red) were genetic structures observed in that sequence. Middle: PCR products for Il6 gene of WT, IL6-DIO-Het, and IL6-DIO-KO mice yielded the expected bands. Right: IL-6 protein in serum was undetectable by ELISA in IL6-DIO-KO mice after LPS administration. All results are represented as mean ± SEM; ★p ≤ 0.05 vs. LPS-C57BL/6 mice