Salmon lice are ectoparasites on salmonids and feed on blood, mucus, and skin from their hosts. This causes high annual costs for treatment and control for the aquaculture industry. Salmon lice have a life cycle consisting of eight life stages. Sex determination by eye is only possible from the sixth stage onwards. A molecular sex determination has not been carried out so far, even though few individual sex-linked SNPs have been reported. In the present study, we used known sex-specific SNPs as a basis to sequence the complete sex-specific gene variants and used the sequence information to develop a sex determination assay. This assay could be used to determine the developmental speed of the two sexes already in the earliest life stages. Additionally, we sampled salmon lice in the nauplius II stage, determined the sex of each individual, pooled their RNA according to their sex, and used RNA sequencing to search for differences in gene expression and further sex-specific SNPs. We succeeded in developing a sex-determination assay that works on DNA or RNA from even the earliest larval stages of the salmon louse after hatching. At these early developmental stages, male salmon lice develop slightly quicker than females. We detected several previously unknown, sex-specific SNPs in our RNA-data seq, but only very few genes showed a differential expression between the sexes. Potential connections between SNPs, gene expression, and development are discussed.