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. 2001 Feb 6;98(4):1889–1894. doi: 10.1073/pnas.041390398

Figure 3.

Figure 3

Nsp1 is an essential factor for transcription but not replication of the EAV genome. (A) Immunoprecipitation analysis of nsp1 (29 kDa) and nsp2 (61 kDa) expression. (B) Northern blot analysis of intracellular RNA (24 h after transfection) with a probe complementary to the 3′ end of all EAV mRNAs (10). The positions of the EAV genome and six sg mRNAs are indicated. The band labeled E in the DITRAC lane represents an extra sg RNA transcribed from a cryptic TRS in the EMCV IRES (see text). (C) RT-PCR analysis of sg-RNA7 synthesis. The generation of minus- (Upper) or plus-stranded (Lower) sg RNA7 in transfected cells was tested by RNA7-specific RT-PCR. One of the primers was located in the RNA7 body and the other in the leader sequence, thereby generating an RNA7-specific PCR product of 540 bp (arrowhead). Mutant EAV030F, which generates about 500-fold reduced levels of sg RNAs (17, 20), was included to confirm the sensitivity of the assay.