ANAT 262 Lecture Notes - Lecture 12: Differential Centrifugation, Randy Schekman, Secretion
15 Jul 2016
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Document Summary
Much of our knowledge of important proteins in the secretory pathway comes from the use of yeast genetics in the lab of randy schekman. Amazingly, most of this work was found to be valid also in mammalian cells. Yeasts exposed to mutagen and grow at 24c (can secrete since normal t). It was assumed cells would increase in density (more protein) if they could not secrete. Shifted to 37c for 3 hours, extra-heavy yeast separated by density centrifugation. Moved back to 24c (assumption of secretion block continued too long it would kill yeast) Mutations in 23 genes identified in first screen, more later. Initial isolation of mutants required density gradient centrifugation: two major centrifugation techniques. Differential: achieved by subjecting a suspension of cellular components (obtained by disrupting the cells by homogenization), to the action of centrifugal forces. The sedimentation coefficient depends on the shape and size of the particles and organelles.
