Fetal Whole Genome Sequencing from Chorionic Villi

Genomic DNA was isolated from chorionic villi or tissue using a TIANamp Genomic DNA Kit (Beijing Tiangen Biotech Co. Ltd., Beijing, China). Genomic DNA from POCs was sheared to 250 to 300 bp fragments using an Ion Shear Plus Reagents Kit (Thermo Fisher Scientific, Waltham, MA). Ion Torrent barcoded libraries were made using an Ion Plus Fragment Library Kit (Thermo Fisher Scientific). An Ion PGM Template OT2 200 Kit (Thermo Fisher Scientific) was used for template amplification and enrichment of the target sequence. Ion sphere particles were recovered, and template-positive ion sphere particles were enriched using an Ion OneTouch ES (Thermo Fisher Scientific). Sequencing was performed using an Ion PGM Sequencing 200 Kit v2 (Thermo Fisher Scientific) on a 318 sequencing chip for a total of 500 nucleotide flows, yielding average read lengths of 220 to 230 bp. Ten DNA samples were pooled together and labeled with different barcodes on the 318 chip. The average whole genomic sequence depth was approximately 0.02×, and the average read number was approximately 500 K. The primary sequencing BAM data were submitted to the PGX cloud server (available at http://www.pgxcloud.com/), which was offered by a third-party company (JBRH, China), to analyze the chromosomal CNVs. The data analysis pipeline was established according to previous reports.^{[12 (link),13 (link)]}