Protocol for measuring the auxin-induced changes of m⁶A modification

Area of Science:

• Plant molecular biology
• Epigenetics
• RNA biology

Background:

• N6-methyladenosine (m⁶A) is a dynamic RNA modification in plants.
• Plant hormones are key regulators of these m⁶A dynamics.
• Understanding these regulatory mechanisms is crucial for plant development.

Purpose of the Study:

• To present a detailed protocol for detecting phytohormone-induced changes in plant m⁶A modification.
• To provide a reproducible method for analyzing m⁶A dynamics in response to plant hormones.
• To facilitate further research into hormone-mediated epigenetic regulation in plants.

Main Methods:

• The protocol involves auxin treatment of plant samples.
• RNA extraction followed by RNA dot blot assays.
• Preparation of m⁶A sequencing (m⁶A-seq) libraries.
• Bioinformatic analysis of sequencing data.

Main Results:

• The described protocol enables the detection of changes in m⁶A levels.
• It allows for the analysis of m⁶A modification dynamics induced by specific plant hormones, such as auxin.
• The method is suitable for both qualitative and quantitative assessments of m⁶A changes.

Conclusions:

• This protocol offers a robust approach for studying plant hormone-regulated m⁶A modifications.
• It can be applied to investigate the roles of various phytohormones in modulating m⁶A patterns.
• The findings contribute to a deeper understanding of post-transcriptional gene regulation in plants.