# Quantitative Analysis of Sex-Specific Feminizer (fem) Transcripts During Honey Bee (Apis mellifera) Development

## TL;DR

This study quantifies sex-specific fem transcripts in honey bees across developmental stages, revealing differences in expression between males and females.

## Contribution

The study introduces optimized Real-Time PCR methods to accurately quantify femF and femM transcripts in honey bees.

## Key findings

- femF is 100-fold more expressed in females than in males.
- femM shows only 10-fold higher expression in males compared to females.
- femM transcripts in males are stable and not degraded despite a premature stop codon.

## Abstract

Sex determination in honey bees (Apis mellifera) is controlled by the complementary sex determiner (csd) gene, which directs female- or male-specific splicing of the downstream feminizer (fem) transcript. Previous studies have reported contradictory data on the expression of fem transcripts in both sexes, but no rigorous quantitative analysis across developmental stages had been performed. Here, we optimized Real-Time PCR conditions to reliably detect and quantify both female-specific (femF) and male-specific (femM) transcripts, addressing challenges posed by AT-rich sequences, repeated regions, and cDNA instability. Using these methods, we analyzed transcript levels in eggs, larvae, and pupae of both sexes. Our results show that femF is highly specific for females, with approximately 100-fold higher expression in females than in males, whereas femM is less sex-specific, with only ~10-fold higher expression in males even at early developmental stages. Notably, femF transcripts are detectable in males, and femM expression increases in females during later pupal stages. Quantitative comparison indicates that femM expression in males is similar to femF expression in females, indicating that despite the presence of the premature stop codon in the male transcript, this transcript is not degraded through the mRNA surveillance mechanism. Our study provides a framework for evaluating fem transcript dynamics and has important implications for interpreting sex-determination mechanisms in honey bees.