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De Novo Transcriptome Sequencing of the Snail Echinolittorina malaccana: Identification of Genes Responsive to Thermal Stress and Development of Genetic Markers for Population Studies

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Abstract

Echinolittorina snails inhabit the upper intertidal rocky shore and face strong selection pressures from thermal extremes and fluctuations. Revealing the molecular processes of adaptive significance is greatly obstructed by the scarcity of genomic resource for these taxa. Here, we reported the first comprehensive transcriptome dataset for the genus Echinolittorina. Using Illumina HiSeq 2000 platform, about 52 M and 54 M paired-end clean reads were, respectively, generated for the control and heat-stressed libraries. Totally, 115,211 unique transcript fragments (unigenes) were assembled, with an average length of 453 bp and a N50 size of 492 bp. Approximately one third of the unigenes could be annotated according to their homology matches against the Nr, Swiss-Prot, COG, or KEGG databases, and they were found to represent 23,098 non-redundant genes. Gene expression comparison revealed that 1,267 and 6,663 annotated genes were, respectively, up- and downregulated with at least twofold changes upon heat stress. Gene Ontology and KEGG pathway analyses indicated that there were overrepresented amount of genes enriched in a broad spectrum of biological processes and pathways, including those associated with cytoskeleton organization, developmental regulation, signaling transduction, infection, and cardiac function. In addition, a transcriptome-wide search for polymorphic loci yielded a total of 11,228 simple sequence repeats (SSRs) from 9,938 unigenes and 138,631 single nucleotide polymorphism (SNP) and insertion/deletion (INDEL) sites among 22,770 unigenes. The large number of transcript sequences acquired, the biological pathways identified, and the candidate microsatellite and SNP/INDEL loci discovered in the study will serve as valuable resources for further investigations of genetic differentiation and thermal adaptation among populations.

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Acknowledgments

We are grateful to two anonymous reviewers for their comments on an earlier version of this manuscript. The authors thank Aldrin KY Yim for the access of server at the Hong Kong Bioinformatics Centre, The Chinese University of Hong Kong (CUHK) and Long Fan for his assistance in compiling the PERL scripts to process the data. The project was supported by a grant of the Group Research Scheme from the Research Committee, CUHK.

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  1. School of Life Sciences, The Chinese University of Hong Kong, Shatin, Hong Kong

    Wei Wang, Jerome H. L. Hui, Ting Fung Chan & Ka Hou Chu

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  1. Wei Wang
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Correspondence to Ka Hou Chu.

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Figure S1

E-value and similarity distribution in Nr annotation. (A) 54.6 % of the annotated unigenes displayed strong homology (e-value smaller than 1.0E-15) to known genes, whereas 45.4 % of the sequences have e-values ranging from 1.0E-15 to 1.0E-5. (B) 63.9 % of the unigenes showed a similarity higher than 40 %, and 36.1 % of the sequences had a similarity ranging from 40 % to 15 %. (JPEG 77 kb)

Table S1

Statistics of functional annotation. The number of unigenes annotated to each database is shown. (PDF 80 kb)

Table S2

Annotation of the assembled sequences. The unigenes were analyzed for similarities with sequences recorded in major databases, including the nr, Swiss-Pro, COG and KEGG databases. (XLSX 14567 kb)

Table S3

KEGG pathways present in the snail transcriptome. A total of 23,016 unigenes could find homology hits in the KEGG database. These unigenes represent 258 predicted KEGG pathways. The number of unigenes in each pathway is summarized. (XLSX 20 kb)

Table S4

List of unigenes differentially expressed upon heat stress. A total of 8,304 transcript-derived unigenes were detected as up-regulated while 22,359 unigenes were down-regulated. Annotation of these unigenes to the nr database is provided. (XLSX 3456 kb)

Table S5

List of over-represented KEGG pathways. 51 KEGG pathways were found significantly over-represented in the heat-stressed animals. For each pathway, the number of unigenes differentially expressed, the total number of unigenes, the corrected p-value and pathway ID are shown. (PDF 9 kb)

Table S6

List of SNP/INDEL loci identified in the assembled unigenes. A total of 138,631 candidate SNP/INDEL loci were identified among 22,770 unigenes. (XLSX 11143 kb)

Table S7

List of protein-coding unigenes with SNP/INDEL sites. 8,590 unigenes with SNP/INDEL sites corresponded to protein-coding genes. Annotation of these unigenes is provided. (XLSX 774 kb)

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Wang, W., Hui, J.H.L., Chan, T.F. et al. De Novo Transcriptome Sequencing of the Snail Echinolittorina malaccana: Identification of Genes Responsive to Thermal Stress and Development of Genetic Markers for Population Studies. Mar Biotechnol 16, 547–559 (2014). https://doi.org/10.1007/s10126-014-9573-0

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