Identification of copy number variations and common deletion polymorphisms in cattle

Abstract

Background: Recently, the discovery of copy number variation (CNV) led researchers to think that there are more variations of genomic DNA than initially believed. Moreover, a certain CNV region has been found to be associated with the onset of diseases. Therefore, CNV is now known as an important genomic variation in biological mechanisms. However, most CNV studies have only involved the human genome. The study of CNV involving other animals, including cattle, is severely lacking.

Results: In our study of cattle, we used Illumina BovineSNP50 BeadChip (54,001 markers) to obtain each marker's signal intensity (Log R ratio) and allelic intensity (B allele frequency), which led to our discovery of 855 bovine CNVs from 265 cows. For these animals, the average number of CNVs was 3.2, average size was 149.8 kb, and median size was 171.5 kb. Taking into consideration some overlapping regions among the identified bovine CNVs, 368 unique CNV regions were detected. Among them, there were 76 common CNVRs with > 1% CNV frequency. Together, these CNVRs contained 538 genes. Heritability errors of 156 bovine pedigrees and comparative pairwise analyses were analyzed to detect 448 common deletion polymorphisms. Identified variations in this study were successfully validated using visual examination of the genoplot image, Mendelian inconsistency, another CNV identification program, and quantitative PCR.

Conclusions: In this study, we describe a map of bovine CNVs and provide important resources for future bovine genome research. This result will contribute to animal breeding and protection from diseases with the aid of genomic information.

Figure 1

Map of identified copy number variations in Bos taurus coreanae. The locations of all copy number variation regions (CNVRs) are depicted by triangles (red color: gain; blue color: loss). The thick line (color: green) indicates common CNVRs (freq. > 2.5%).

Figure 2

Visualization and validation of copy number variation region (chr15:1836732-2039483) by visual examination and quantitative PCR. (A) Visualization of identified individual copy number variations in UCSC Genome Browser. The black bars indicate copy number variation of each sample. (B) Determination of copy number by quantitative PCR around third marker (Hapmap24310-BTA-162764). (C) Visual examination by consecutive genoplot images of markers. The first marker shows a monomorphic pattern having 2× (color: blue), 1× (color: cyan), and 0× (color: black). The samples having a deletion (copy number: 1×; color:cyan) were consecutively displayed in deletion position to the fourth marker.

Figure 3

Distribution of identified common deletion polymorphisms according to the frequency of parent (sire)--child (steer) heritability (P-C) error.

Figure 4

Scheme of identification of common deletion polymorphisms by parent (sire)--child (steer) heritability (P-C) error and validation by quantitative PCR. (A) Scheme of genoplot image in Illumina BeadStudio 3.2 software where SNP marker was located within copy number variation region. Steer and sire were marked as "X" and "O", respectively. The three dotted lines represent three SNP genotypes (A/A, A/B, and B/B). (B) Genoplot image showing P-C heritability error. The steer (child) is shown on the left side (copy number: 1×; CNV genotype: A/-; marked at "X") and its sire (parent) on the right (copy number: 1×; CNV genotype: B/-; marked at "O"). (C) Difference of SNP and CNV genotype in one pedigree. (D) Heritability error table. Nine P-C errors in one marker are displayed. The table shows the sample ID of steer (child) and sire (parent) and their SNP genotypes having heritability error. (E) Genoplot image of identified common deletion polymorphisms (marker name: ARS-BFGL-NGS-24778). Three types of copy number (2×, 1×, and 0×) are depicted. Individuals having hemizygous deletions (copy number: 1×) clustered into two distinct groups (color: cyan). Samples having null copy number are displayed with a black dot at the bottom. (F) Validation by qPCR around the ARS-BFGL-NGS-24778 marker. The individuals having homozygous (null) and hemizygous deletions in the genoplot image were spotted approximately in the same copy number position by qPCR.