Lecture 23 - Lecture 23: Molecular techniques Gene...

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Lecture 23: Molecular techniques Gene targeting
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ene targeting Gene targeting Transgene -> nock out - Stable incorporation of gene isruption of specific gene Knock out > Knock in -> Disruption of specific gene Replacement of specific gene Dominant negative -> Incorporation of dominant mutant allele causing the same phenotype as loss-of-function RNA interference, knock down -> Depletion of mRNA
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Gene knock out in yeast Based on high efficiency of homologous recombination esign of “disruption construct” (PCR) Design of disruption construct (PCR) antibiotic (e.g. kanamycin) resistance gene lectable marker = selectable marker
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Gene knock out in yeast Integration of kanMX ene in chromosome gene in chromosome
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Gene knock out in yeast Systematic disruption of 6000 genes in yeast -> 4500 not required for viability Conditional knock out ? 1) Deletion gene A 2) Transformation with vector containing gene A under control of GAL1 promoter -> medium with galactose: gene expressed medium with glucose: gene not expressed
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Conditional knock out 1) Deletion gene A 2) Transformation with vector containing gene A under control of GAL1 romoter medium with galactose: gene expressed promoter -> medium with glucose: gene not expressed wild type A - A - + Gal1-A + galactose + glucose non viable viable viable non viable (if A is an essential gene)
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This note was uploaded on 11/23/2010 for the course MATH math 203 taught by Professor Dr.who during the Spring '10 term at University of Toronto- Toronto.

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Lecture 23 - Lecture 23: Molecular techniques Gene...

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