215-Lab4A-Spotting_Assay

215-Lab4A-Spotting_Assay - Lab 4A Set up Spotting Silencing...

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Lab 4A –Set up Spotting Silencing Assays 4A-1 Lab 4A Transcriptional Silencing Spotting Silencing Assays In this experiment, each person will perform a silencing spot assay to quantitatively assay the level of silencing of the hml::TRP1 reporter. In this experiment you will plate dilutions of 1) a blank plasmid (pRS425); 2) a plasmid with the wild type Hst::Sir2 chimera (pJM447); and 3-6) four of your mutant plasmids to test if your Hst:Sir chimera mutants can silence transcription. 1. Label 30 microcentrifuge tubes RS-1 to RS-5, JM-1 to JM-5, M1-1 to M1-5, M2-1-M2-5 . Tubes RS-1 to RS-5 will contain dilutions of the strain with plasmid pRS425 Tubes JM-1 to JM-5 will contain dilutions of the strain with plasmid pJM477 Tubes M1-1 to M1-5 will contain dilutions of one of the mutant strains (e.g. T2JS1.09). Tubes M2-1 to M2-5 will contain dilutions of one of the mutant strains (e.g. T2JS2.09). Tubes M3-1 to M3-5 will contain dilutions of one of the mutant strains (e.g. T2JS4.09). Tubes M4-1 to M4-5 will contain dilutions of one of the mutant strains (e.g. T2JS6.09). 2.
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215-Lab4A-Spotting_Assay - Lab 4A Set up Spotting Silencing...

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