Truseq nano dna library

Whole genome sequencing (WGS) to a depth of 30X coverage was performed using Illumina HiSeq X sequencer for N=126 IMPACT and N=120 POISED samples. The samples for WGS were prepared to the Illumina TruSeq Nano DNA library or TruSeq DNA PCR-free library preparation guides. Assembly of each individual genome was performed using the Isaac aligner (9 (link)). The DRAGEN Germline Small Variant Caller was used to call both SNVs and small indels, and to yield a genome variant file (gVCF) that includes variants along with quality metrics. We performed sample-based QC and dropped two samples because of sex inconsistencies from the POISED dataset. We also filtered variants with GQX < 30, DP < 7, SNP hard quality < 10.41, low depth DP <= 1, ploidy conflict, and variants not meeting thresholds for median base quality of alternate reads and likelihood.
We used the WGS data to impute HLA alleles using the HISAT-genotype software that utilizes HISAT2 (hierarchical indexing for spliced alignment of transcripts 2) alignment system to align DNA sequences using a graph Ferragina Manzini index (10 (link)). We had high quality (call rate) of imputation for HLA-DQA1*01:02 allele in both IMPACT (98.81%) and POISED (98.75%).

DNA libraries were prepared from tumor and matched normal tissue (blood) available for five pediatric meningioma cases (three with recurrent tumor samples available) and sequenced on three lanes each on HiSeq2000 instruments (2 × 100 bp). DNA libraries were prepared according to the Illumina TruSeq Nano DNA Library protocol using the TruSeq DNA Nano kit (Illumina, Hayward, CA) and sequenced on one lane on HiSeq X (2 × 151 bp) using the HiSeq X Ten Reagent Kit v2.5 (both Illumina).

WGS was performed in the two murine cell lines MM5080 and MM9275 and the corresponding matched germline DNAs. Briefly, genomic DNA was purified using a NucleoSpin Tissue kit (Macherey-Nagel). DNA quality and concentration were evaluated with a Qubit System (Invitrogen). Next-generation sequencing capture libraries were prepared according to the TruSeq Nano DNA Library (Illumina) and were sequenced using a 150 base-pair paired-end read protocol by Macrogen on an Illumina NovaSeq 6000. The resulting FASTQ file analysis was performed by the Genome One platform (Dreamgenics) using the HMMcopy adaptation of CopywriteR.