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Long undecoded transcript isoform (LUTI) detection in meiotic budding yeast by direct RNA and transcript leader
Amy Tresenrider1,2, Minghao Chia3,4, Folkert J van Werven4
1Department of Molecular and Cell Biology, Barker Hall, University of California, Berkeley, Berkeley, CA 94720, USA.
STAR Protocols
|February 16, 2022
Summary
This study identifies meiosis-specific Long Undecoded Transcript Isoforms (LUTIs) in yeast using direct RNA sequencing and TL-seq. These findings help understand how LUTIs regulate gene expression during meiosis.
Area of Science:
- Molecular Biology
- Genetics
- Yeast Biology
Background:
- Long Undecoded Transcript Isoforms (LUTIs) are 5'-extended mRNAs.
- LUTIs are poorly translated and can downregulate proximal promoter transcription.
- Understanding LUTI roles in gene regulation is crucial.
Purpose of the Study:
- To systematically identify meiosis-specific LUTIs in budding yeast.
- To quantify the abundance of LUTIs and canonical transcripts.
- To provide a protocol for LUTI identification and quantification.
Main Methods:
- PolyA RNA extraction from synchronized meiotic yeast cells.
- Long-read direct RNA sequencing.
- Transcript leader sequencing (TL-seq) for LUTI identification and quantification.
Main Results:
- Identification of meiosis-specific LUTIs.
- Quantification of LUTI and gene-proximal (PROX) transcript abundance.
- Demonstration of TL-seq's utility in studying LUTIs.
Conclusions:
- A robust protocol for identifying and quantifying LUTIs during meiosis is established.
- This method enables systematic study of LUTI function in gene regulation.
- Findings contribute to understanding mRNA regulation in yeast meiosis.
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