Skip to main content
SpringerLink
Log in

De novo assembly, gene annotation, and molecular marker development using Illumina paired-end transcriptome sequencing in the clam Saxidomus purpuratus

  • Research Article
  • Published:
Genes & Genomics Aims and scope Submit manuscript

Abstract

The clam Saxidomus purpuratus is an important economic marine bivalve species in China. In this study, we performed a de novo transcriptome sequencing of S. purpuratus gill tissues to generate a transcriptome dataset by performing Illumina paired-end sequencing on the HiSeq 2500 platform. A total of 10,488,024 raw reads were produced. After removing the inferior sequences, adaptor sequences, and rRNAs, 68,080,636 high-quality reads were obtained, from which 5,115,494 assembled contigs were produced. These contigs were assembled into 120,479 transcripts and 66,388 unigenes with mean lengths of 674.96 and 562.70 bp, respectively. All unigene sequences were compared against the NCBI databases, and 26,781 unigenes were annotated. Both the gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG) pathway analysis showed that several functional genes were involved in the stress response, immune system, and oxidation reduction process. Meanwhile, 414 simple sequence repeats (SSRs) were identified in the S. purpuratus transcriptome. SSR validation via PCR confirmed that 26 primer pairs had expected products, and 13 primers proved to be polymorphism among 30 individuals. The S. purpuratus transcriptome advances the underlying molecular understanding of this economic shellfish and provides a basis for further exploring S. purpuratus genomics resources.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  • Altschul SF, Madden TL, Schäffer AA, Zhang JH, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • An HS, Lee JW (2012) Development of microsatellite markers for the Korean mussel, mytilus coruscus (Mytilidae) using next-generation sequencing. Int J Mol Sci 13:10583–10593

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Anestis A, Pörtner HO, Lazou A, Michaelidis B (2008) Metabolic and molecular stress responses of sublittoral bearded horse musselModiolus barbatusto warming sea water: implications for vertical zonation. J Exp Biol 211:2889–2898

    Article  CAS  PubMed  Google Scholar 

  • Apweiler R, Bairoch A, Wu CH, Barker WC, Boeckmann B, Ferro S, Gasteiger E, Huang HZ, Lopenz R, Magrane M, Martin MJ, Natale DA, O’Donovan C, Redaschi N, Yeh LSL (2004) UniProt: the universal protein knowledgebase. Nucleic Acids Res 32:D115–D119

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ashburner M, Ball CA, Blake JA, Botstein D, Butler H, Michael-Cherry J, Davis AP, Dolinski K, Dwight SS, Eppig JT (2000) Gene ontology: tool for the unification of biology. Nat Genet 25:25–29

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bi JH, Li Q, Zhang XJ, Zhang ZX, Tian JL, Xu YS, Liu WG (2016) Seasonal variation of biochemical components in clam (Saxidomus purpuratus Sowerby 1852) in relation to its reproductive cycle and the environmental condition of Sanggou Bay, China. J Ocean U China 15:341–350

    Article  CAS  Google Scholar 

  • Cho ES, Seo YI, Suh YS (2013) Genetic analysis of the purplish Washington clam (Saxidomus purpuratus Sowerby) of Korean coastal waters. J Environ Biol 34:613–621

    CAS  PubMed  Google Scholar 

  • Choi JW, Kim SK, Choi YS, Ryu TK (2003) Optimal conditions for artificial fertilization, embryonic development, and larval growth of the purple clam, Saxidomus Purpuratus from southern coast of Korea. Korean J Malacol 19:33–40

    Google Scholar 

  • Choi YK, Jo PG, Choi CY (2008) Cadmium affects the expression of heat shock protein 90 and metallothionein mRNA in the pacific oyster,Crassostrea gigas. Comp Biochem Physiol C Toxicol Pharmacol 147:286–292

    Article  PubMed  Google Scholar 

  • Clark MS, Thorne MAS, Vieira FA, Cardoso JCR, Power DM, Peck LS (2010) Insights into shell deposition in the Antarctic bivalve Laternula elliptica: gene discovery in the mantle transcriptome using 454 pyrosequencing. BMC Genomics 11:362

    Article  PubMed  PubMed Central  Google Scholar 

  • Conesa A, Götz S, García-Gómez JM, Terol J, Talón M, Robles M (2005) Blast GO: a universal tool for annotation, visualization and analysis in functional genomics research. Bioinformatics 21:3674–3676

    Article  CAS  PubMed  Google Scholar 

  • Deng YY, Li JQ, Wu SF, Zhu YP, Chen YW, He FC (2006) Integrated nr database in protein annotation system and its localization. Comput Eng 32:71–74

    Google Scholar 

  • Deng YW, Lei QN, Tian QL, Xie SH, Du XD, Li JH, Wang LQ, Xiong YX (2014) De novo assembly, gene annotation, and simple sequence repeat marker development using Illumina paired-end transcriptome sequences in the pearl oyster Pinctada maxima. Biosci Biotech Bioch 78:1–8

    Article  Google Scholar 

  • Dimitriadis VK, Gougoula C, Anestis A, Pörtner HO, Michaelidis B (2012) Monitoring the biochemical and cellular responses of marine bivalves during thermal stress by using biomarkers. Mar Environ Res 73:70–77

    Article  CAS  PubMed  Google Scholar 

  • Feng BB, Dong LL, Niu DH, Meng SS, Zhang B, Niu DB, Hu SN, Li JL (2010) Identification of immune genes of the Agamaki clam (Sinonovacula constricta) by sequencing and bioinformatic analysis of ESTs. Mar Biotechnol 12:282–291

    Article  CAS  PubMed  Google Scholar 

  • Fu DK, Chen JH, Zhang Y, Yu ZN (2011) Cloning and expression of a heat shock protein (HSP) 90 gene in the haemocytes ofCrassostrea hongkongensisunder osmotic stress and bacterial challenge. Fish Shellfish Immunol 31:118–125

    Article  CAS  PubMed  Google Scholar 

  • Grabherr MG, Haas BJ, Yassour M, Levin JZ, Thompson DA, Amit I, Adiconis X, Fan L, Raychowdhury R, Zeng QD et al (2011) Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nat Biotechnol 29:644–652

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kanehisa M, Goto S, Kawashima S, Hattori M (2004) The KEGG resource for deciphering the genome. Nucleic Acids Res 32:D277–D280

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kim YH, Chang DS, Cha BY (2001) Catch yield fluctuation and relative growth of the purplish Washington clam Saxidomus purpuratus (Sowerby) in the South Sea, Korea. Fisheries. Aquat Sci 4:98–100

    Google Scholar 

  • Kim SK, Rosenthal H, Clemmesen C, Park KY, Kim DH, Choi YS, Seo HC (2005) Various methods to determine the gonadal development and spawning season of the purplish Washington clam, Saxidomus purpuratus (Sowerby). J Appl Ichthyol 21:101–106

    Article  Google Scholar 

  • Kim YH, Ryu DK, Lee DW, Chang DS, Kim JB, Kim ST, Kwon DH (2006) Morphological analysis among populations of purpulish Washington clam, Saxidomus purpuratus on the Korean waters. Korean J Malacol 22:23–26

    CAS  Google Scholar 

  • Li HJ, Liu SX, He CB, Gao XG, Yuan XT (2013) Identification of a small HSP gene from hard clam Meretrix meretrix and its potential as an environmental stress biomarker. Aquat Biol 18(3):243–252

    Article  CAS  Google Scholar 

  • Li HJ, Zhang JJ, Li HB, Gao XG, He CB (2014) Development and characterization of 13 polymorphic microsatellite markers for the Chinese surf clam (Mactra chinensis) through Illumina paired-end sequencing. Conserv Genet Resour 6:877–879

    Article  Google Scholar 

  • Niu DH, Wang L, Sun FY, Liu ZJ, Li JL (2013) Development of molecular resources for an intertidal clam, Sinonovacula constricta, using 454 transcriptome sequencing. PloS ONE 8:e67456

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Novaes E, Drost DR, Farmerie WG, Pappas-Jr GJ, Grattapaglia D, Sederoff RR, Kirst M (2008) High-throughput gene and SNP discovery in Eucalyptus grandis, an uncharacterized genome. BMC Genomics 9:312

    Article  PubMed  PubMed Central  Google Scholar 

  • Núñez-Acuña G, Gallardo-Escárate C (2013) Identification of immune-related SNPs in the transcriptome of Mytilus chilensis through high-throughput sequencing. Fish Shellfish Immunol 35:1899–1905

    Article  PubMed  Google Scholar 

  • Pauletto M, Milan M, Moreira R, Novoa B, Figureras A, Babbucci M, Patarnello T, Bargelloni L (2014) Deep transcriptome sequencing of Pecten maximus hemocytes: a genomic resource for bivalve immunology. Fish Shellfish Immunol 37:154–165

    Article  CAS  PubMed  Google Scholar 

  • Snyder MJ (2000) Cytochrome P450 enzymes in aquatic invertebrates: recent advances and future directions. Aquat Toxicol 48(4):529–547

    Article  CAS  PubMed  Google Scholar 

  • Tatusov RL, Galperin MY, Natale DA, Koonin EV (2000) The COG database: a tool for genome-scale analysis of protein functions and evolution. Nucleic Acids Res 28:33–36

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wang Y, Wang AM, Guo XM (2010) Development and characterization of polymorphic microsatellite markers for the Northern Quahog Mercenaria mercenaria (Linnaeus, 1758). J Shellfish Res 29:77–82

    Article  Google Scholar 

  • Wang HX, Huan P, Lu Xia, Liu BZ (2011) Mining of EST-SSR markers in clam Meretrix meretrix larvae from 454 shotgun transcriptome. Genes Genet Syst 86:197–205

    Article  CAS  PubMed  Google Scholar 

  • Wang RJ, Li C, Stoeckel J, Moyer G, Liu ZJ, Peatman E (2012) Rapid development of molecular resources for a freshwater mussel, Villosa lienosa (Bivalvia: Unionidae), using an RNA-seq-based approach. Freshw Sci 31:695–708

    Article  CAS  Google Scholar 

  • Wei LP, Shu YF, Guan FT, Han YP, Yu ZR (1982) A preliminary survey on the biology of Saxidomus purpuratus. J Fisheries China 6:1–8 (Chinese edition with English abstract)

    Google Scholar 

  • Wei LP, Wang BG, Sun YT, Yang FS, Wang SF (1994) Study on resource enhancement technology of Saxidomus purpuratus. Fish Sci Technol Info 21:13–18 (Chinese edition with English abstract)

    Google Scholar 

  • Wu HB, Gong H, Liu P, He XL, Luo SB, Zheng XM, Zhang CY, He XM, Luo JN (2014) Large-scale development of EST-SSR markers in sponge gourd via transcriptome sequencing. Mol Breed 34:1903–1915

    Article  CAS  Google Scholar 

  • Yoon JM, Park SY (2006) Genetic differences and variation in two purple Washington clam (Saxidomus purpuratus) populations from south and north Korea. Korean J Malacol 22:97–108

    Google Scholar 

  • Yue X, Liu BZ, Sun L, Tang BJ (2011) Cloning and characterization of a HSP70 gene from Asiatic hard clamMeretrix meretrixwhich is involved in the immune response against bacterial infection. Fish Shellfish Immunol 30:791–799

    Article  CAS  PubMed  Google Scholar 

  • Zhang LL, Li L, Zhang GF (2011) A Crassostrea gigas Toll-like receptor and comparative analysis of TLR pathway in invertebrates. Fish Shellfish Immun 30:653–660

    Article  CAS  Google Scholar 

  • Zhang JJ, Li HJ, Qin YJ, Ye S, Liu M (2016) Identification of functional genes involved in Cd2+ response of Chinese surf clam (Mactra chinensis) through transcriptome sequencing. Environ Toxicol Pharmacol 41:113–120

    Article  CAS  PubMed  Google Scholar 

  • Zhao XX, Wang QH, Jiao Y, Huang RL, Deng YW, Wang H, Du XD (2012) Identification of genes potentially related to biomineralization and immunity by transcriptome analysis of pearl sac in pearl oyster Pinctada martensii. Mar Biotechnol 14:730–739

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This study was supported by Grants from National Natural Science Foundation of China (Grant Nos. 31572595).

Author information

Authors and Affiliations

  1. National Marine Environmental Monitoring Center, Dalian, 116023, Liaoning, People’s Republic of China

    Hongjun Li, Min Liu & Sheng Ye

  2. Dalian Ocean University, Dalian, 116023, Liaoning, People’s Republic of China

    Min Liu, Sheng Ye & Feng Yang

Authors
  1. Hongjun Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Min Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Sheng Ye
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Feng Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Hongjun Li.

Ethics declarations

Conflict of interest

Hongjun Li declares that he has no conflict of interest. Min Liu declares that she has no conflict of interest. Sheng Ye declares that he has no conflict of interest. Feng Yang declares that she has no conflict of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (XLSX 94 KB)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Li, H., Liu, M., Ye, S. et al. De novo assembly, gene annotation, and molecular marker development using Illumina paired-end transcriptome sequencing in the clam Saxidomus purpuratus . Genes Genom 39, 675–685 (2017). https://doi.org/10.1007/s13258-017-0535-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s13258-017-0535-6

Keywords

Search

Navigation