2D PAGE: Sample Preparation and Fractionation by Anton Posch

By Anton Posch

This publication provides wide insurance of the foundations and up to date advancements of pattern instruction and fractionation instruments in Expression Proteomics as a rule and two-dimensional electrophoresis (2-DE) specifically. With its certain potential to solve hundreds of thousands of proteins in one run, 2-DE remains to be a basic study device for almost all protein-related medical projects.

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Mutanolysin (Sigma M-9901). Protease inhibitor cocktail (Sigma P8465). 0. 1 M NaCl, protease inhibitor cocktail. 6. 2 % (v/v) Triton X-100. 2. Yeast Lysis 1. Purified yeast lytic -1,3-endoglucanase. A recombinant endoglucanase is recommended (BGLII from C. ) is also available. 28 Salazar 2. S. cerevisiae strain rhSOD 2060 411 SGA122 (MAT , leu2; Chiron Corporation) (other strains used with similar results are: INVSc‘1 (his3 1, leu2 trp1-289 ura352; Invitrogen); DSY-5 (MATa leu2 trp1 ura3-52 his3 pep4 prb1; Dualsystems Biotech).

0. 1 M NaCl, protease inhibitor cocktail. 6. 2 % (v/v) Triton X-100. 2. Yeast Lysis 1. Purified yeast lytic -1,3-endoglucanase. A recombinant endoglucanase is recommended (BGLII from C. ) is also available. 28 Salazar 2. S. cerevisiae strain rhSOD 2060 411 SGA122 (MAT , leu2; Chiron Corporation) (other strains used with similar results are: INVSc‘1 (his3 1, leu2 trp1-289 ura352; Invitrogen); DSY-5 (MATa leu2 trp1 ura3-52 his3 pep4 prb1; Dualsystems Biotech). 3. YPD: 10 g/L yeast extract, 20 g/L tryptone, 2 % (w/v) glucose.

A. (1986) Isolation of rat intestinal microsomes: partial characterization of mucosal cytochrome P-450. Arch Biochem Biophys 244, 492–501. 6. Nally, J. , Whitelegge, J. , Pereira, M. , Blanco, D. , and Lovett, M. A. (2005) Purification and proteomic analysis of outer membrane vesicles from a clinical isolate of Leptospira interrogans serovar Copenhageni. Proteomics 5, 144–152. 7. , and Dallner, G. (1982) Fractionation of isolated liver cells after disruption with a nitrogen bomb and sonication.

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