Journal: Nutrition & Metabolism
Article Title: Short-term arginine deprivation results in large-scale modulation of hepatic gene expression in both normal and tumor cells: microarray bioinformatic analysis
Figure Lengend Snippet: RT-PCR confirmation of selected rat hepatic gene expression in normal and tumor cells in response to 18 hr arginine deprivation . Semi-quantitative Reverse Transcription Polymerase Chain Reaction was used to assess relative expression of rat LDLr, farnesyl diphosphate synthase (FDPS), GADD45, Insig-1, and GAPDH for cells cultured with (+) or without (-) arginine, Adult liver served as a positive control. DNase treated total RNA was reverse-transcribed with and without (negative control) reverse transcriptase. A . Electrophoresis of PCR products was performed in 2% agarose, 1× TAE gel and visualized by ethidium bromide staining; 1 kb DNA marker was used to verify size of the PCR products. B . Densitometry of gel bands was assessed via LabWorks Software and values were ratioed to GAPDH to provide relative comparisons between Arginine + and Arginine - conditions.
Article Snippet: DNase treated total RNA was reverse-transcribed with and without reverse transcriptase for 60 minutes at 50 C using Superscript III Reverse Transcriptase (Invitrogen, Carlsbad, CA), Oligo (dT)12–18 (Invitrogen, Carlsbad, CA), RNase OUT ribonuclease inhibitor (Invitrogen, Carlsbad, CA, and 10 mM DNTP set (Invitrogen, Carlsbad, CA.
Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Cell Culture, Positive Control, Negative Control, Electrophoresis, Polymerase Chain Reaction, Staining, Marker, Software