TSS identification of Rap1-regulated transcripts by 5' end RNA sequencing

Eukaryotic cells utilize several mechanisms to ensure that expression of aberrant non-coding RNAs is limited. Gene looping, chromatin modification or remodeling, and RNA surveillance contribute to ensure the fidelity of transcription and limit non-coding transcripts. We have identified that the transcription factor Rap1 is critical for limiting the expression of aberrant RNAs, particularly near the highly expressed ribosomal protein genes. Here we utilize a 5' end RNA sequencing protocol to identify transcript TSSs at single nucleotide resolution in wild-type and Rap1 auxin-inducible degron strains of Saccharomyces cerevisiae. Overall design: 5' end RNA sequencing analysis of Saccharomyces cerevisiae S288C strains transformed with Os TIR1 ligase and auxin-inducible degron (IAA7) tagged Rap1, or wild-type control strains. 3 biological replicates of each condition are analysed. For each biological replicate, cells were grown overnight in YPD medium, diluted, and treated with 500 µM 3-indole-acetid acid (3-IAA, auxin) for RAP1_AID_IAA samples or left untreated for WT samples. RAP1_AID_IAA samples were taken 2 hours after treatment, and for WT control samples were taken in mid-logarithmic growth. Details for library preparation are described below.

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Wu ACK, Patel H, Chia M, Moretto F, Frith D, Snijders AP, van Werven FJ