access quick kit rt‐pcr system  (Promega)

Journal: PLoS ONE

Article Title: A High Diversity of Eurasian Lineage Low Pathogenicity Avian Influenza A Viruses Circulate among Wild Birds Sampled in Egypt

doi: 10.1371/journal.pone.0068522

Figure Lengend Snippet: Number of influenza A virus PCR positive specimen and number of embryonated chicken egg isolates.

Article Snippet: The surface and internal protein genes were then amplified using influenza A virus specific primers as overlapping fragments with the Access Quick one-step RT-PCR kit (Promega, Madison, WI) and subsequently sequenced on an automated Applied Biosystems 3730 system using cycle sequencing dye terminator chemistry (Life Technologies, Carlsbad, CA).

Techniques: Isolation, Northern Blot

Journal: Molecular & Cellular Proteomics : MCP

Article Title: Reductive Stress Selectively Disrupts Collagen Homeostasis and Modifies Growth Factor-independent Signaling Through the MAPK/Akt Pathway in Human Dermal Fibroblasts

doi: 10.1074/mcp.RA118.001140

Figure Lengend Snippet: PDGF signaling and induction of an ER stress response in BJ Fibroblasts. A, Lysates from BJ fibroblasts treated with 10 ng ml−1 PDGF-BB for 10′ or 24 h in serum free media were analyzed by 10% SDS-PAGE followed by immunoblotting for p-PDGFR, p-p44/42 and eIF4E. PDGF-BB initiated a signaling response within 10′. A representative image of n = 3 biological replicates is shown; kDa markers are shown on the left. B, Schematic RT-PCR assay illustrating the expected PCR products of XBP1 depending on splicing/activation status. When XBP1 is activated by ER stress, the PstI restriction site is removed and PstI digestion no longer occurs. C, RT-PCR products from BJ fibroblasts treated ± 10 ng ml−1 PDGF-BB ± DTT were analyzed by 2% agarose gel electrophoresis following digestion with PstI for the expression of XBP1 (upper panel) and β Actin (lower panel). DTT resulted in the splicing of XBP1. bp markers are shown on the left. D, Lysates from BJ fibroblasts treated ± 10 ng ml−1 PDGF-BB for 24 h in serum free media ± DTT were analyzed by 10% SDS-PAGE followed by immunoblotting for p-eIF2α (upper panel) and actin (lower panel). DTT, but not PDGF-BB alone, stimulated the phosphorylation of eIF2α. kDa markers are shown on the left. E, Relative quantitation of the p-eIF2α levels shown in (D)

Article Snippet: RNA content was measured using an ND-1000 spectrophotometer (Nanodrop® Technologies Inc) and 50 ng RNA was subjected to RT-PCR, using the Access-Quick RT-PCR kit (Promega).

Techniques: SDS Page, Western Blot, Reverse Transcription Polymerase Chain Reaction, Activation Assay, Agarose Gel Electrophoresis, Expressing, Quantitation Assay