WGS of Acipenser baerii: 10X Genomics technology
DNA was extracted using a phenol/chloroform protocol and gDNA quality and purity was assessed using spectrophotometry, fluorometry and capillary electrophoresis. 10X Chromium Library was prepared according to 10X Genomics protocols using the Genome Reagent Kits v2. Optimal performance has been characterized on input gDNA with a mean length greater than 40 kb (~158Kb). The libraries were prepared from 0,625 ng of HMW DNA. Briefly, in the microfluidic Genome Chip, a library of Genome Gel Beads is combined with HMW template gDNA in Master Mix and partitioning oil to create Gel Bead-In-EMulsions (GEMs) in the Chromium. Each Gel Bead is functionalized with millions of copies of a 10x Barcoded primer. Upon dissolution of the Genome Gel Bead in the GEM, primers containing (i) an Illumina R1 sequence (Read 1 sequencing primer), (ii) a 16 bp 10x Barcode, and (iii) a 6 bp random primer sequence are released. Read 1 sequence and the 10x Barcode are added to the molecules during the GEM incubation. P5 and P7 primers, Read 2, and Sample Index are added during library construction. 10 cycles of PCR were applied to amplify libraries. Library quality was assessed using a Fragment Analyser and quantified by QPCR using the Kapa Library Quantification Kit. Librray was sequenced on an Illumina HiSeq4000 using a paired-end read length of 2x150 pb with the Illumina HiSeq4000 sequencing kits.