Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, AB T6G 2P5, Canada.
BMC Genomics. 2011 Nov 15;12:559. doi: 10.1186/1471-2164-12-559.
One of the goals of livestock genomics research is to identify the genetic differences responsible for variation in phenotypic traits, particularly those of economic importance. Characterizing the genetic variation in livestock species is an important step towards linking genes or genomic regions with phenotypes. The completion of the bovine genome sequence and recent advances in DNA sequencing technology allow for in-depth characterization of the genetic variations present in cattle. Here we describe the whole-genome resequencing of two Bos taurus bulls from distinct breeds for the purpose of identifying and annotating novel forms of genetic variation in cattle.
The genomes of a Black Angus bull and a Holstein bull were sequenced to 22-fold and 19-fold coverage, respectively, using the ABI SOLiD system. Comparisons of the sequences with the Btau4.0 reference assembly yielded 7 million single nucleotide polymorphisms (SNPs), 24% of which were identified in both animals. Of the total SNPs found in Holstein, Black Angus, and in both animals, 81%, 81%, and 75% respectively are novel. In-depth annotations of the data identified more than 16 thousand distinct non-synonymous SNPs (85% novel) between the two datasets. Alignments between the SNP-altered proteins and orthologues from numerous species indicate that many of the SNPs alter well-conserved amino acids. Several SNPs predicted to create or remove stop codons were also found. A comparison between the sequencing SNPs and genotyping results from the BovineHD high-density genotyping chip indicates a detection rate of 91% for homozygous SNPs and 81% for heterozygous SNPs. The false positive rate is estimated to be about 2% for both the Black Angus and Holstein SNP sets, based on follow-up genotyping of 422 and 427 SNPs, respectively. Comparisons of read depth between the two bulls along the reference assembly identified 790 putative copy-number variations (CNVs). Ten randomly selected CNVs, five genic and five non-genic, were successfully validated using quantitative real-time PCR. The CNVs are enriched for immune system genes and include genes that may contribute to lactation capacity. The majority of the CNVs (69%) were detected as regions with higher abundance in the Holstein bull.
Substantial genetic differences exist between the Black Angus and Holstein animals sequenced in this work and the Hereford reference sequence, and some of this variation is predicted to affect evolutionarily conserved amino acids or gene copy number. The deeply annotated SNPs and CNVs identified in this resequencing study can serve as useful genetic tools, and as candidates in searches for phenotype-altering DNA differences.
家畜基因组学研究的目标之一是识别导致表型性状变异的遗传差异，特别是那些具有经济重要性的性状。描述家畜物种的遗传变异是将基因或基因组区域与表型联系起来的重要步骤。牛基因组序列的完成和 DNA 测序技术的最新进展使得深入研究牛中存在的遗传变异成为可能。在这里，我们描述了两个不同品种的肉牛公牛的全基因组重测序，目的是鉴定和注释牛中的新型遗传变异形式。
使用 ABI SOLiD 系统，将一头黑安格斯公牛和一头荷斯坦公牛的基因组分别测序到 22 倍和 19 倍覆盖率。与 Btau4.0 参考组装的序列比较产生了 700 万个单核苷酸多态性（SNP），其中 24%在两个动物中都有发现。在荷斯坦、黑安格斯和两个动物中发现的总 SNP 中，分别有 81%、81%和 75%是新的。对数据的深入注释鉴定了两个数据集之间超过 16000 个不同的非同义 SNP（85%是新的）。将 SNP 改变的蛋白质与来自许多物种的同源物进行比对表明，许多 SNP 改变了保守的氨基酸。还发现了一些预测会产生或去除终止密码子的 SNP。将测序 SNP 与 BovineHD 高密度基因分型芯片的基因分型结果进行比较，结果表明对于纯合 SNP 的检测率为 91%，对于杂合 SNP 的检测率为 81%。基于对 422 个和 427 个 SNP 的后续基因分型，估计黑安格斯和荷斯坦 SNP 集的假阳性率分别约为 2%。沿着参考组装比较两个公牛的读取深度，鉴定了 790 个可能的拷贝数变异（CNV）。使用实时定量 PCR 成功验证了随机选择的 10 个 CNV，其中 5 个是基因内的，5 个是非基因内的。CNV 富集了免疫系统基因，包括可能有助于泌乳能力的基因。大多数 CNV（69%）在荷斯坦公牛中被检测为丰度较高的区域。
在这项工作中测序的黑安格斯和荷斯坦动物与赫里福德参考序列之间存在大量遗传差异，其中一些变异预计会影响进化保守的氨基酸或基因拷贝数。在重测序研究中鉴定的深度注释 SNP 和 CNV 可用作有用的遗传工具，并作为搜索表型改变 DNA 差异的候选物。
