Transcriptome analysis of sex-related genes in the blood clam Tegillarca granosa

Abstract

BackgroundBlood clams (Tegillarca granosa) are one of the most commercial shellfish in China and South Asia with wide distribution in Indo-Pacific tropical to temperate estuaries. However, recent data indicate a decline in the germplasm of this species. Furthermore, the molecular mechanisms underpinning reproductive regulation remain unclear and information regarding genetic diversity is limited. Understanding the reproductive biology of shellfish is important in interpreting their embryology development, reproduction and population structure. Transcriptome sequencing (RNA-seq) rapidly obtains genetic sequence information from almost all transcripts of a particular tissue and currently represents the most prevalent and effective method for constructing genetic expression profiles.ResultsNon-reference RNA-seq, an Illumina HiSeq2500 Solexa system, and de novo assembly were used to construct a gonadal expression profile of the blood clam. A total of 63.75 Gb of clean data, with at least 89.46% of Quality30 (Q30), were generated which was then combined into 214,440 transcripts and 125,673 unigenes with a mean length of 1,122.63 and 781.30 base pairs (bp). In total, 27,325 genes were annotated by comparison with public databases. Of these, 2,140 and 2,070 differentially expressed genes (DEGs) were obtained (T05 T08 vs T01 T02 T04, T06 T07 vs T01 T02 T04; in which T01-T04 and T05-T08 represent biological replicates of individual female and male clams, respectively) and classified into two groups according to the evaluation of biological replicates. Then 35 DEGs and 5 sex-related unigenes, in other similar species, were investigated using qRT-PCR, the results of which were confirmed to data arising from RNA-seq. Among the DEGs, sex-related genes were identified, including forkhead box L2 (Foxl2), sex determining region Y-box (Sox), beta-catenin (β-catenin), chromobox homolog (CBX) and Sex-lethal (Sxl). In addition, 6,283 simple sequence repeats (SSRs) and 614,710 single nucleotide polymorphisms (SNPs) were identified from the RNA-seq results.ConclusionsThis study provided the first complete gonadal transcriptome data for the blood clam and allowed us to search many aspects of gene sequence information, not limited to gender. This data will improve our understanding of the transcriptomics and reproductive biology of the blood clam. Furthermore, molecular markers such as SSRs and SNPs will be useful in the analysis of genetic evolution, bulked segregant analysis (BSA) and genome-wide association studies (GWAS). Our transcriptome data will therefore provide important genetic information for the breeding and conservation of germplasm.

Gonad transcriptome of golden musselLimnoperna fortuneireveals potential sex differentiation genes

“…According to phylogenetic analysis, the L. fortunei Sox9 grouped with other proteins from the group E, showing closer relation to the bivalve sequences (Sox8 and Sox9 from C. gigas and P. fucata, respectively) with high bootstrap support (100). Consistent with other studies done with mollusks (11,23,25), Lf-Sox9 did not show male-biased expression, as the expression of this transcript was similar in males and females, i.e., unbiased expressed. This suggests that Sox9 gene may not be related to sex differentiation mechanisms in the golden mussel.…”

“…FoxL2 gene has been found in several other bivalves such as C. farreri, Pinctada margaritifera,H. shlegelii,Crassostrea hongkongensis,Tegillarca granosa, and Chlamys nobilis, all showing a sexually dimorphic pattern of expression towards females (14)(15)(16)20,21,23). However, in C. gigas gonad transcriptome, FoxL2 was expressed in both sexes, it was highly but not specifically expressed in the ovary, with increased expression earlier during sexual development in females.…”

“…Previous research on sex-determining pathway genes has focused on homologous sequences of model organisms. However, identification and expression of genes related to these processes have been studied in some bivalves, including the vertebrate female-determining gene forkhead box L2 (FoxL2) (19,20,14,21), the male-determining genes double-sex-and mab-3-related transcription factor (Dmrt) (18,20,21), and Sox (Sryrelated HMG box) family genes (11,12,15,16,22,23). FoxL2 and Dmrt are the key genes for sex determination and maintenance, respectively, and their antagonic role in the control of gonadal sex has been characterized in mammals and has been suggested to play similar antagonic roles in bivalve species (12,24).…”

“…FoxL2 and Dmrt are the key genes for sex determination and maintenance, respectively, and their antagonic role in the control of gonadal sex has been characterized in mammals and has been suggested to play similar antagonic roles in bivalve species (12,24). Some of those aforementioned genes usually present sex-biased expression in bivalve gonads, such as SoxH/Sox30 (11,12,22) and Dmrt-like (14,15,23) are male-based and FoxL2 is female-biased (11,14,16,(20)(21)(22)(23). Additionally, other conserved sex-determining related genes from model species have been identified in bivalves, such as Wnt4 (wingless-type MMTV integration site family, member 4 (11,15,16), -catenin) (11,15,16,25), and Rspo1 (R-spondin1) (15,16), among others.…”

Transcriptomics in aquaculture: current status and applications

“…To conserve its germplasm resource, Chen et al . () analysed the gonadal gene expression and identified many sex related genes. Mun et al .…”