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Analysis of Myc-Tagged Proteins in Cyc8, Tup1, and Gcn4 Mutants via Western Blot

This study investigates the expression levels of Myc-tagged proteins such as Spt7p, Srb6p, TBP, Rpb3p, Cyc8p, and Tup1p in various mutant strains, including cyc8∆, tup1∆, and gcn4∆. Utilizing Western blot analysis, we assess the expression patterns of these proteins under specific growth conditions. Whole cell extracts (WCEs) from independent cultures are analyzed, employing anti-myc and anti-Gcd6p antibodies for robust detection. Our findings will enhance the understanding of protein interactions within these mutant backgrounds, shedding light on their functional roles.

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Analysis of Myc-Tagged Proteins in Cyc8, Tup1, and Gcn4 Mutants via Western Blot

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  1. A cyc8∆ tup1∆ WT gcn4∆ h. c. GCN4-HA h. c. GCN4-HA h. c. GCN4-HA vector vector vector vector myc-Spt7p Gcd6p B cyc8∆ tup1∆ WT gcn4∆ h. c. GCN4-HA h. c. GCN4-HA h. c. GCN4-HA vector vector vector vector myc-Srb6p Gcd6p cyc8∆ tup1∆ WT gcn4∆ h. c. GCN4-HA h. c. GCN4-HA h. c. GCN4-HA vector vector vector vector myc-TBP Gcd6p D cyc8∆ tup1∆ WT gcn4∆ h. c. GCN4-HA h. c. GCN4-HA h. c. GCN4-HA vector vector vector vector myc-Rpb3p Gcd6p E GCN4 TUP1-myc CYC8-myc WT gcn4∆ tup1∆ WT gcn4∆ cyc8∆ myc-Cyc8p myc-Tup1p Gcd6p CYC8-myc TUP1-myc h. c. GCN4-HA WT gcn4∆ tup1∆ WT gcn4∆ cyc8∆ myc-Tup1p FIG. S1. Western analysis of myc-Spt7p, myc-Srb6p, myc-TBP, myc-Rpb3, myc-Cyc8p and myc-Tup1p expression in cyc8∆, tup1∆ and gcn4∆ mutants. (A-F) myc-tagged strains used in Figs. 5 and 7 were grown under the same conditions used for ChIP analysis. WCEs were prepared and subjected to Western analysis using anti-myc or anti-Gcd6p antibodies as a loading control. Two independent cultures of each strain were analyzed in successive lanes of the gel. C F myc-Cyc8p Gcd6p Fig. S1

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