Exploring valid reference genes for quantitative real-time PCR analysis in Sesamia inferens (Lepidoptera: Noctuidae)

Area of Science:

• Molecular Biology
• Entomology
• Pest Management

Background:

• The pink stem borer (Sesamia inferens) is a significant rice pest in Asia, causing substantial yield losses.
• Gene expression studies in S. inferens are limited, hindering research into its biology and control.
• Quantitative real-time PCR (qRT-PCR) requires reliable reference genes for accurate gene expression normalization.

Purpose of the Study:

• To evaluate the suitability of seven candidate reference genes for normalizing gene expression in Sesamia inferens.
• To identify optimal reference genes across different experimental conditions, including tissues, developmental stages, sex, and temperatures.
• To validate the selected reference genes using a target gene, heat shock protein 83 (hsp83).

Main Methods:

• Seven candidate reference genes were assessed: 18S rRNA, EF1, GAPDH, RPS13, RPS20, TUB, and ACTB.
• Gene expression stability was analyzed using qRT-PCR under various conditions (tissues, developmental stages, sex, temperature).
• The expression of hsp83 was analyzed using the identified optimal reference genes for validation.

Main Results:

• Ribosomal protein S13 (RPS13), ribosomal protein S20 (RPS20), and elongation factor 1 (EF1) were optimal for normalizing gene expression in different S. inferens tissues.
• 18S ribosomal RNA (18S rRNA), EF1, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were best for normalization across developmental stages and sex.
• 18S rRNA, RPS20, and tubulin (TUB) were optimal for fifth instar larvae exposed to varying temperatures.

Conclusions:

• This study provides validated reference gene sets for accurate gene expression analysis in Sesamia inferens.
• The identified reference genes are crucial for future research on S. inferens gene expression under diverse experimental conditions.
• These findings will aid in a deeper understanding of the pink stem borer's biology and the development of effective pest management strategies.