A Comparative Analysis Of Transcriptions Of Huaxi Crab And Non - Tooth - Concealed

Brachyura are known as crabs. There are about 32 percent of all known 6,973 brachyura species occupied freshwater habitats (Ng et al,2008, Samuel et al,2011). Most of them habitat in the sea in their whole lifecycle, a few species live in freshwater and salt marshes, but they are not fully cast off the yoke of the sea. When laying eggs, they must be returned to the sea, such as Eriocheir spp., these crabs are also known as adults limnic non-marine crabs (Schubart et al,2000). Other groups are fully adapted to the freshwater environment. They are considered as truly freshwater crab, such as Sinopotamon spp. in China inland. However, lacking of genome data limit the research of freshwater adaptation of crabs. Transcriptome, as a set of all protein coding gene, can provide valueable data. The emerging of next-generation sequencing technologies provided a cost-efficiency approach for high-throughput sequence determination, and advantages to accelerate de novo transcriptome sequencing for organisms of unknown genomic sequences.In the present research, two transcriptome libraries, sampled by pooling liver, gill, muscle and antenna gland,from Sinopotamon yangtsekiense and Sesarma neglectum were constructed. A total amount of 59,084,458 and 62,386,654 raw reads were generated using Illumina sequencing platform. After removing the low quality reads, de novo assembly was carried out with Trinity program. We obtained 95,148 for S.yangtsekiense and 102,019 for S. neglectum unigenes, respectively. More than 30 percent unigenes were annotated based on sequence similarities with several protein databases using Blast. Blasted with the GO and COG databases, unigenes were classified into different functional categories. Additionally, abundant simple sequence repeats were also detected. Overall, our results greatly enriched the transcriptome information of brachyura.We identified 4,903 pairs of orthologous genes between their transcriptomes using bidirectional best hit method and estimated their rates of nonsynonymous (Ka) and synonymous (Ks) substitutions. Among them,7 orthologous gene pairs have a Ka/Ks rate between 0.5 and 1, and 2 orthologous gene pairs have a Ka/Ks value larger than 1. Based on RPKM method, we gained 121,218 significantly differentially expressed genes between transcriptomes.These genes were further blasted with GO database and KEGG pathway database. By comparing with the DGE database of Eriocheir sinensis japonica witch sampled separately at different development stages, 5 genes witch related to freshwater adaptation were identified. They are V-type proton ATPase, voltage-dependent anion-selective channel, heat shock protein 70, heat shock protein 90 and cytoplasmic carbonic anydrase. We verified expressing changing of these 5 gene pairs in condition of exposed into different salinity in the gills of Sinopotamon yangtsekiens and Sesarma neglectum using semiquantitative RT-PCR, providing an important resource for further research.