gene screen plus membranes  (New England Nuclear Corporation)

Journal: Gene Expression

Article Title: Interferon regulatory factor-1 (IRF-1) activates the synthetic IRF-1-responsive sequence (GAAAGT) 4 in Saccharomyces cerevisiae

doi:

Figure Lengend Snippet: IRF-1 transcript level in uninduced and galactose-induced diploid yeast cells containing an IRF-1 expression plasmid. A–C: Diploid cells contained the IRF-1 expression plasmid YEpGIR and the YEpUAS reporter (Fig. 1B) with (A) VREαl, (B) VREβ”, (C) (GAAAGT)4. In the control sample (D), the diploid cells contained Z669 and YGRA2. Cells were grown for 24–38 hours in galactose (GAL) or glucose (GLU) medium, and the RNA extracted according to Vijayraghavan et al. (1989). Samples (5 μg) were run through a 1 % agarose gel. Endogenous ribosomal RNA was used as molecular weight marker. The samples were transferred to Gene Screen nylon membranes (DuPont) and hybridized as described (Büeler et al., 1992). The probe was the 1141 bp Nco I fragment of IRF-1 cDNA (Miyamoto et al., 1988) labeled by random priming (Feinberg and Vogelstein, 1983) and used at 3.5 × 105 cpm/ml. Autoradiography was for 1 hour.

Article Snippet: The samples were transferred to Gene Screen nylon membranes (DuPont) and hybridized as described ( Büeler et al., 1992 ).

Techniques: Expressing, Plasmid Preparation, Control, Agarose Gel Electrophoresis, Molecular Weight, Marker, Labeling, Autoradiography