Transcriptomic analysis of genes involved in reproduction at different ages in Daphnia pulex (Branchiopoda, Cladocera)

Daphnia pulex is a freshwater microcrustacean that is known for its cyclical parthenogenesis. In D. pulex, parthenogenetic reproduction switches to sexual reproduction when the living conditions worsen. However, this transformation also occurs over age under favourable living conditions. Thus, there might be a relationship between aging and reproductive conversion. We performed Illumina RNA sequencing, generating 51 712 680 and 59 854 588 raw reads from 1 day-old D. pulex and 25 day-old D. pulex, respectively. From these reads, 60 776 transcripts were assembled and 36 569 (60.15%) unigenes were annotated. A number of significantly differentially expressed genes associated with growth, aging, and reproduction were identified and Quantitative real-time PCR for six genes confirmed the transcriptome data. RNA interference (RNAi) of the caspase-3 gene (casp3) that is a key gene for growth, development, aging, and reproduction, was conducted, which achieved an 80% reduction in casp3 mRNA expression. Meanwhile, the mRNA expression of upstream genes jnk and akt, and sir2 (encoding a substrate of casp3) were also detected. The change of jnk mRNA expression before and after RNAi was not significant (P>0.05) and the mRNA levels of sir2 decreased, while akt increased after casp3 RNAi (P<0.05). The results indicated the interrelationships of some genes in the senescence pathway and helped to identify the molecular mechanism of the aging progress in D. pulex. Overall, the difference in mRNA expression profile during aging of D. pulex forms a basis for further studies aimed at understanding the role of the transcriptional level in regulating aging and reproductive transformation.