The genome assembly, annotations, and all raw data associated with this project are affiliated with NCBI BioProject PRJNA914702.
The organism used to generate this assembly was a male red sea urchin, samples ID "Mf1", with a 101 mm test diameter. DNA for all sequencing was extracted from gonad tissue following a modified CTAB extraction procedure (please reference publication for additional information).
Dissections were done by Andrea Bodnar, and DNA extractions by Jennifer Polinski.
DNA was mechanically sheared to ~500 bp on a Covaris M220a and bead-cleaned/size-selected with PCRclean DX beads (Aline Biosciences) prior to library prep following manufacturer's SOP with the KAPA HyprePrep DNA kit (Roche). 1.5 ug of DNA went into the KAPA prep, a double-sides size selection targeting fragments 450-600 bp was done, and no PCR amplification steps were performed. The resulting library was sequenced at GMGI on a NextSeq 500 using a v2.5 high-output 2x150 kit.
Illumina library prep and sequencing were done by Jennifer Polinski.
Prior to ONT library prep, DNA was size-selected for large fragments using SPRI beads with custom buffers (buffer 1 = ONT "SPRI size selection protocol for >1.5-2 kb" & Stortchevoi et al. 2020). DNA size distribution and quantity were determined using the Fragment Analyzer gDNA kit (Agilent) and Qubit dsDNA BR assay. 1 ug of HMW DNA was prepared for ONT sequencing with the NAnopore Ligation Sequencing Kit (SQK-LSK109) following manufacturer's specifications, and sequenced on a MinION sequencers. Four ONT library preps and sequencing runs were performed to achieve desired coverage.Basecaling was performed using ONT Guppy with the following command: ont-guppy-cpu/bin/guppy_basecaller -i ./fast5/ -s ./fastq/ -c /data/app/nanopore/ont-guppy-cpu/data/dna_r9.4.1_450bps_hac.cfg --num_callers 10.
ONT library prep and sequencing were done by Jennifer Polinski.
Hi-C libraries were generated by Phase Genome using gonad tissue from Mf1.
Information for each step of genome assembly and analysis can be found in the appropriate directory here, in the order listed.