Comparing control options for time-series RNA sequencing experiments in nonmodel organisms: An example from grasses

Area of Science:

• Plant Science
• Molecular Biology
• Genomics

Background:

• RNA sequencing (RNA-seq) is crucial for studying gene expression, especially in time-course experiments.
• Two control strategies exist: temporal control (TC) and biological control (BC).
• Direct comparisons of TC and BC effects on differential gene expression analysis are lacking.

Purpose of the Study:

• To compare the impact of temporal control (TC) versus biological control (BC) on time-course RNA-seq analyses.
• To evaluate the effects of control type on identifying differentially expressed genes (DEGs) and functional annotations.
• To assess these impacts in drought stress experiments on two wild grass species (Paspalum).

Main Methods:

• Performed time-course RNA-seq experiments on two Paspalum species under drought stress.
• Utilized both temporal control (TC) and biological control (BC) strategies.
• Analyzed and compared the number of loci, differentially expressed genes (DEGs), and enriched Gene Ontology terms between control types.

Main Results:

• Biological control (BC) assemblies yielded a higher number of loci in both species.
• DEG numbers increased with stress and decreased at recovery for both TC and BC.
• High correlation (r = 0.653–0.852) in DEG expression levels and similar functional annotation results were observed between control types.

Conclusions:

• Both temporal control (TC) and biological control (BC) provide comparable results for differential gene expression and functional annotation in time-course RNA-seq.
• The optimal control choice depends on experimental specifics such as duration, organism, cost, and labor.
• This study provides valuable insights for researchers designing time-course transcriptomic experiments.