Genomic resources of two landsnail, Aegista diversifamilia and Dolicheulota formosensis, generated by Illumina paired-end sequencing

Introduction

Stylommatophora is the dominant gastropod clade on land. The family Bradybaenidae is mainly distributed in Asia and diversified in East Asia. However, only a handful of genetic markers are available for phylogenetic research. We selected two land snail species as representatives of Bradybaenidae: Aegista diversifamilia Huang et al., 2014 and Dolicheulota formosensis (H. Adams, 1866). We subjected these species to Illumina paired-end sequencing in the hope that this will expand and provide valuable genetic resources for phylogeographic and phylogenetic research of gastropods.

Materials and methods

Genomic sequences were generated from one fresh specimen preserved at -20°C (A. diversifamilia) and one two-year-old ethanol-preserved specimen (D. formosensis), respectively. The living individual of A. diversifamilia was collected from leaf litter under lowland broadleaf forest in Xiulin Township, Hualien County, Taiwan in October 2013. D. formosensis was collected from the tree trunk of a lowland broadleaf forest in Mudan Township, Pingtung County, Taiwan in June 2012. Snails were brought back to laboratory and rinsed with ddH₂O to eliminate soil and other particles attached to the shell. The living individual of A. diversifamilia was preserved in a -20°C refrigerator. The individual of D. formosensis was relaxed under water for 12 hours and fixed by blanching with boiling water for several seconds. The specimen of D. formosensis was transferred to 95% ethanol solution and the ethanol was changed every 24 hours for 48 hours. The specimen of D. formosensis was finally preserved in 95% ethanol.

For A. diversifamilia, the shell was removed after the snail was crushed. Soft tissue was rinsed with TEK buffer (50 ml 1M Tris-HCl pH 7.5, 50 ml 0.2M EDTA pH 7.5, 15g KCl, pH 7.5) and then used for genomic DNA extraction with AxyPrep™ Multisource Genomic DNA Miniprep Kit (Axygen Bioscience) following the manufacturer’s protocol. For D. formosensis, partial foot tissue was rinsed with ddH₂O several times to remove ethanol and then used for DNA extraction through a modified phenol-chloroform method (Jiang et al., 1997).

The quantity of extracted DNA solution was evaluated by Qubit^® dsDNA HS Assay Kit (Life Technologies). The DNA concentration was 131 ng/μL (total mass 20.96 μg) and 52.9 ng/μL (5.92 μg) for A. diversifamilia and D. formosensis, respectively. The DNA solution was send to BGI (Shenzhen, China) for paired-end library construction with insert size of approximately 500 base pairs using a Paired-End DNA Sample Prep Kit (Illumina). Sequencing was performed using Illumina HiSeq2000 in BGI. Raw sequences data can be accessed in Sequence Read Archive under accession number SRR1918809 for A. diversifamilia and SRR1920140 for D. formosensis.

Ethics policies

Ethical approval for the animals used was not required. Both species used in this study were common and not under list of protected species in Taiwan.