Genome Sequencing of Deep-Sea Hydrothermal Vent Worm

The samples of “long-skinny” morphotype of R. piscesae were obtained during Alvin dive 4243 from the deep-sea hydrothermal vent at Cathedral vent, Main Endeavor Field of the Juan de Fuca Ridge (47° 56’ N, 129° 05’ W, 2,181 m depth) on August 9, 2006. Genomic DNA (gDNA) was extracted from vestimentum muscle of the specimen using a standard phenol/chloroform extraction protocol and broken into random fragments for whole-genome shotgun (WGS) sequencing. Agarose gel electrophoresis was used to check the quality of the gDNA, and Qubit system was used to quantify the gDNA. Short-insert paired-end libraries (180 bp, 300 bp and 500 bp) were prepared using the NEBNext Ultra DNA Library Prep Kit for Illumina (NEB, USA) according to the standard protocol, respectively. Large-insert mate-pair libraries (2 kb, 5 kb, 10 kb and 15 kb) were prepared following the Cre-lox recombination-based protocol [64 (link)]. All DNA libraries were subjected to paired-end sequencing on the Illumina Hiseq 2000 platform (Illumina). Muscle samples from vestimentum were also collected for constructing RNA sequencing (RNA-seq) library. Total RNA was extracted with TRIzol reagent (Molecular Research Center, USA). Paired-end library for RNA-seq was constructed using the Paired-End Sample Preparation Kit (Illumina Inc., San Diego, CA, USA) and sequenced on the Illumina Hiseq 2000 platform (Illumina).