Author: Zirkel, Florian; Kurth, Andreas; Quan, Phenix-Lan; Briese, Thomas; Ellerbrok, Heinz; Pauli, Georg; Leendertz, Fabian H.; Lipkin, W. Ian; Ziebuhr, John; Drosten, Christian; Junglen, Sandra
Title: An Insect Nidovirus Emerging from a Primary Tropical Rainforest Document date: 2011_6_14
ID: ulwo6i38_29
Snippet: Identification of subgenomic mRNAs. For Northern blotting, the Northern Blot Starter Kit (Roche, Mannheim, Germany) was used. Digoxigenin (DIG)-labeled probes were generated by PCR using the primers shown in Fig. 4B and listed in Table S1 in the supplemental material. Total RNA of isolate CAVV/C79/CI/2004 was extracted with the Qiagen RNeasy Kit (Qiagen, Hilden, Germany) from C6/36 cells at 24 hpi. RNA was separated on a 2% formaldehyde-1.5% agar.....
Document: Identification of subgenomic mRNAs. For Northern blotting, the Northern Blot Starter Kit (Roche, Mannheim, Germany) was used. Digoxigenin (DIG)-labeled probes were generated by PCR using the primers shown in Fig. 4B and listed in Table S1 in the supplemental material. Total RNA of isolate CAVV/C79/CI/2004 was extracted with the Qiagen RNeasy Kit (Qiagen, Hilden, Germany) from C6/36 cells at 24 hpi. RNA was separated on a 2% formaldehyde-1.5% agarose gel, blotted onto a nylon membrane (Roche, Mannheim, Germany), and hybridized with the CAVV-specific, DIG-labeled probes. RNAs were analyzed by chemiluminescence using 1:10,000 anti-DIG-alkaline phosphatase Fab fragments and 1:100 CDP-Star reagent (Roche, Mannheim, Germany).
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