Impaired inflammasome activation and bacterial clearance in G6PD deficiency due to defective NOX/p38 MAPK/AP-1 redox signaling

Yen, Wei-Chen; Wu, Yi-Hsuan; Wu, Chih-Ching; Lin, Hsin-Ru; Stern, Arnold; Chen, Shih-Hsiang; Shu, Jwu-Ching; Tsun-Yee Chiu, Daniel

The total RNA was extracted using TRIzol Reagent (Life Technologies, CA, USA). RNA was reverse transcribed into cDNA by oligo-dT (Bioman Scientific, Taipei, Taiwan) as the primer in the presence of reverse transcriptase (Superscript III, Invitrogen). qRT-PCR was conducted using a SsoFast™ EvaGreen® Supermix reagent (Bio-Rad, CA, USA) with an iQ5 real-time thermal cycler (Bio-Rad, CA, USA). The expression levels were normalized to those of endogenous ACTB, and the data were analyzed using the 2 −ΔΔCt method. The sequences of the primers used in qRT-PCR are as follows: IL1B: 5′-TGTCTGGTCCATAT GAACTG-3′ and 5′-GCTGTAGAGTGGGCTTATC-3'; ACTB: 5′-TCCACCT TCCAGCAGATG-3′ and 5′-GTGTAACGCAACTAAGTCATAG-3'.