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Protocol publication

[…] DNA degradation and contamination were monitored on 1% agarose gels and the concentration and purity were assessed on NanoDrop 2000 (Thermo Scientific Inc. Waltham, DE, USA); the high-quality DNAs were then used for library construction. Two paired-end libraries were constructed for each individual, the read length was 2 × 100 bp, and whole genome sequencing was performed using Illumina Hiseq2000 instruments (Illumina Inc., San Diego, CA, USA). All processes were performed according to the standard manufacturer’s protocols. In order to get high-quality data, we removed low-quality reads and those containing primer/adaptor contamination which existing in the raw sequencing data by utilizing NGS QC Toolkit with default parameters [-l 75 -q 30] []. After data filtering, we used the Burrows-Wheeler Aligner (BWA) program [] with default parameters [-A1 –B4] to perform sequence alignment based on the UMD3.1 genome assembly which was retrieved from the UCSC website (http://genome.ucsc.edu/). To save run time during the downstream analysis, we converted the SAM files to BAM files and then sorted and merged them by SAMtools []., CNVnator was run on merged BAM files with a bin size of 200 bp following the authors’ recommendations []. After calling, quality control was performed on the raw CNVs for each bull. The filtering criteria included P-value <0.01 (pval1 calculated using t-test statistics), size >1 kb, and q0 < 0.5. P-value <0.01 means that the region between two calls is not a same CNV and q0 means fraction of mapped reads with zero quality. In addition, the CNVs that overlapped with gaps or unplaced chromosomes (chrUn in UCSC) were removed., According to the EBVs for PP and FP, the 8 Holstein bulls were divided into 2 groups, high-group and low-group, and the differential CNVs between the high and […]

Pipeline specifications

Software tools NGS QC Toolkit, BWA, SAMtools, CNVnator
Organisms Cucumber necrosis virus, Bos taurus, Homo sapiens