title: Mitgenome sequence of Halichoeres margaritaceus
design: DNA extraction and quality assessment50-100 mg tissue cut up into piece was added to the preheated (56C) 2 ml EP tube loading with 1ml lysis buffer (100L 20mg/mL Proteinase K and 100L 20%SDS) and Incubated at 56C for 60~120min. The tube was Centrifuged at maximum speed for 10 minutesto get the supernatant after cooling to room temperature. The supernatant was transferred to a new 2.0ml tube, adding equal volume of supernatant of Phenol / Chloroform / isoamylalcohol (25: 24:1) and Centrifuged at maximum speed for 10 minutes. Transferring the supernatant to a new 1.5mL tube and adding 2/3th volume of supernatant of isopropyl alcohol (add 1/10th volume of 3M sodium acetate if necessary), then it was inverted at least 3 times and placed at -20C for 2 hours for precipitation. To remove the supernatant, the tube was centrifuged at 18213g for 10 minutes getting the DNA pellet and it was washed with 1 ml 75% ethanol. Re-suspend the pellet by centrifuging at maximum speed for 5 minutes at room temperature and Completely remove the supernatant. Air-dry the DNA pellet in the biosafety cabinet for a few minutes and add 25L~100L of TE Buffer to dissolve the DNA pellet. DNAs Concentration was detected by Qubit Fluorometer. Sample integrity and purity were detected by Agarose Gel Electrophoresis.Library preparation and sequencingDNBSEQ-T71-1.5g genomic DNA was randomly fragmented by Covaris, The fragmented DNA was selected by Agencourt AMPure XP-Medium kit to an average size of 200-400bp. The selected fragments were through end-repair, 3 adenylated, adapters-ligation, PCR Amplifying and the products were recovered by the AxyPrep Mag PCR clean up Kit. The double stranded PCR products were heat denatured and circularized by the splint oligo sequence. The single strand circle DNA (ssCir DNA) were formatted as the final library and qualified by QC. The qualified libraries were sequenced on MGISEQ-T7 platform.Data quality controlTo make sure reads reliable, MGI paired-ended sequenced Raw reads for genomic survey were first filtered using the fastp (v.0.20.0) preprocessor (set to default parameters) to remove low quality reads, adapters, and reads containing poly-N. The low-quality reads were filtered under the following conditions: (1) reads with 10% unidentified nucleotides (N); (2) reads with > 10 nucleotides aligned to the adapter, allowing 10% mismatch; (3) reads with >50% bases having Phred quality < 5; (4) Removing putative PCR duplicates generated by PCR amplification in the library construction process (read 1 and read 2 of two paired-end reads that were completely identical). To check for contamination, 100,000 reads were selected randomly and matched against sequences in the NT (nucleotide sequence database) library.
library_name: D221000187