FIG. 6.

RT-PCR confirmation of array. (A) The induction patterns of representative genes identified as candidates for alternative pathway effectors were validated by RT-PCR. Total RNA was isolated from TD and IFNAR1^−/− BMDC either untreated, IFN-α/β primed (1,000 IU/ml for 6 h), or SB infected (12 h p.i.) and quantitated. cDNA was synthesized by RT from 50 ng of each RNA with dT₍₁₈₎ primers and amplified by PCR with gene-specific primers (Table 1). (B) cDNA was synthesized from 50 ng of total RNA described above by using a murine ZAP-specific antisense primer, and then amplified by PCR with a ZAP-specific primer pair (Table 1). The same primers were used to amplify a product from plasmid DNA encoding murine ZAP, pcDNA4/to/myc-mouseZAP995, to confirm product size. For both panels, PCR products visualized by gel electrophoresis are indicative of the presence of mRNAs in the original sample. β-Actin mRNA was amplified to confirm the presence of equal RNA, and the results from one experiment representative of three are shown.