DOI: 10.5061/dryad.jh9w0vt7x
title: A chromosome-scale reference genome and genome-wide genetic variations elucidate adaptation in yak

description: Yak is an important livestock for the people who lived in harsh and
 oxygen-deprived Qinghai-Tibetan Plateau and Hindu-Kush Himalayan
 Mountains. Although there is a yak genome be sequenced in 2012, the
 assembly is quite fragmented due to the limitation of Illumina sequencing
 technology. An accurate and complete reference genome is critical for
 studying genetic variation of a specie. Long-read sequences are more
 complete than short-read ones, and they have been successfully used for
 high-quality genome assembly in several species. Here, we present a
 high-quality assembly of the yak genome (PB_v1.0) at chromosome scale,
 which was constructed using long-read sequencing technology assisted by
 chromatin interaction technology. Compared to the previous yak genome
 assembly (BosGru_v2.0), the PB_v1.0 assembly has substantially improved
 chromosome sequence continuity, minimized repetitive structure ambiguity,
 and achieved gene model completeness. To intensively characterize genetic
 variation of yak, we generated de novo genome assemblies based on Illumina
 short reads of seven recognized domestic yak breeds from Tibet and Sichuan
 as well as one wild yak from Hoh Xil. By comparing these eight assemblies
 to the PB_v1.0 genome, we obtained a comprehensive map of yak genetic
 diversity at whole genome level and identified a few protein-coding genes
 that were absent from the PB_v1.0 assembly. Although wild yak suffered
 bottleneck effect, the genetic diversity of wild yak is still higher than
 that of domestic yak. By whole genome alignment, we identified
 breed-specific sequences and genes, this will help the breeds
 identification of yak.
description: High-quality DNA was extracted from the peripheral blood of a
 female yak in Riwoqe County, Tibet. SMRT sequencing libraries were
 constructed with a Blood&Cell Culture DNA Mini Kit (Qiagen,
 Hilden, Germany). A total of 142 SMRT cells generated 184.6 Gbp of subread
 bases with a mean read length of 9.5 kbp on a PacBio RS II instrument
 (Pacific Biosciences, Menlo Park, CA, USA). The Falcon (v. 0.5.0) pipeline
 was used for the initial assembly. The first step was to identify all
 overlaps in the raw reads. Then, the read error was corrected by
 leveraging the overlap information. The second step was to detect overlaps
 in the corrected reads. This step required no consensus calling. The final
 step was to generate the string graph assembly and the contig sequence
 output in FASTA format. To improve the quality of the initial assembly,
 113.34 Gbp of Illumina short reads were generated from the same
 individual. Using Pilon(v1.23)8, 845,002 homozygous insertions, 166,908
 deletions, and 2,355,196 substitutions were identified and corrected. DNA
 from the same individual used in the PacBio sequencing was extracted and
 processed according to BioNano Genomics guidelines. The raw data were
 assembled with the BioNano Solve (v. 3.1.00) assembly pipeline (BioNano
 Genomics, San Diego, CA, USA). The combination of this assembly with the
 initial one yielded a superior assembly with a scaffold N50 of 65.67 Mbp
 and a maximum scaffold length of 128.62 Mbp. Hi-C libraries were created
 from yak whole-blood cells, 2–5 million cells were cross-linked and
 digested with the restriction enzyme <i>Hin</i>dIII. The
 sticky ends of all fragments were biotinylated, ligated to each other to
 form chimeric circles, enriched, sheared, and processed into sequencing
 libraries wherein the individual templates were chimeras of the physically
 associated DNA molecules from the original cross-linking. Hi-C reads was
 generated by Illumina Sequencing platform. The paired-end reads were
 uniquely mapped onto the Bionano assembly, classified into 30 groups using
 3d-DNA(20180922) as the final assembly, and referred to as PB_v1.0. The
 exact locations of each scaffold in the 30 groups were based on the
 collinearity between yak and cattle (UMD3.1.1).  Seven
 domestic yak breeds and one wild yak were selected for whole-genome
 sequencing and assembly. DNA was extracted from the ears of the Tibetan
 breeds, the blood of the Sichuan breeds, and the skin of the wild yak from
 Kunlun Spring, Hoh Xil. A whole-genome shotgun strategy and
 next-generation sequencing (NGS) technologies were run on the Illumina
 HiSeq 2500 platform (Illumina, San Diego, CA, USA). Each genome was
 sequenced with a combination of short-insert (180 bp and 500 bp) and
 long-insert (2 kbp and 5 kbp) DNA libraries. SOAPdenovo (v2.04) was used
 to assemble each genome.