analysis (PCA) using EIGENSOFT3.0 (parameter: -i all.evec -c 1:2 -
p HM:CM:CE:JP -x).
Identification of Insertions and Deletions
Gap tolerant alignments to human reference (hg19) were used to
call indels with program BWA using parameters: aln -o 1 -e 63 -i 15
-L -l 31 -k 2 -t 4. Insertions and deletions (indels) were identified
using samtools, with the command lines as following:
samtools mpileup -ugf ref.fa -b bam.list | bcftools view -bvcg - .
bcftools view var.raw.bcf | vcfutils.pl varFilter -D10000. var.flt.vcf.
Filter criteria were $3 reads support and number of indel
supported reads $30% of all reads mapped to the genomic
position. Indels were called as heterozygous if the indel supported
reads were 30–70% of all reads at that position, and homozygous
if they were greater than 70%.
Validation of SNP and Indel Variants
Forty one SNPs and 18 indels were selected randomly for
validation of SNPs and indels. The selected SNP and indels were
genotyped by PCR and Sanger sequencing. Primers for each
selected SNP and indel were designed based on the IR genome;
the detail sequences of each primer pairs were supplied in Table
S4. The polymerase chain reaction (PCR) was performed in a final
volume of 50 ul with 30 cycles at 94uC for 30 s, annealing
temperature (Table S4) for 30 s, and 72uC for 30 s. The PCR
product was purified and sequenced by BGI (BGI-Shenzhen,
Shenzhen 518083, China).
Table S1 Coverage of targeted orthologous genes.
Coverage of 18,594 gene coding regions of human CCDS by
theoretical target region, target orthologous region and sequencing
data from 3 species.
Table S2 Data production. Summary of captured target
sequence coverage for each non-human primate exome and two
Table S3 Indels. Indels of 21 individuals identified through
aligning the exomes to the human reference genome.
Table S4 Sanger sequencing validation result of SNP
We thank Prof. Rasmus Nielsen from UC-Berkley for his suggestions and
comments. We are also indebted to many additional faculty and staff of
BGI-Shenzhen who contributed to this teamwork. For generously
providing chimpanzee samples, we thank Ngamba Island Chimpanzee
Conceived and designed the experiments: XJ HD Xiaoning Wang Jian
Wang Jun Wang HZ YW. Performed the experiments: BF FS L. Sun MZ
Jufang Wang FL XL FJ Xinzhong Wang BL Y. Zhang JH Jing Wang XZ
MHS. Analyzed the data: XJ MH BF YM LO JR JS L. Song Y. Zhu XF
GZ. Wrote the paper: XJ BF TM MHS HD. Prepared samples: BF FS L.
Sun MZ Jufang Wang FL XL FJ Xinzhong Wang BL Y. Zhang JH Jing
Wang XZ MH. Assisted in writing manuscript: XJ MH BF YM LO JR
TM FS L. Sun JS MZ L. Song Jufang Wang FL Y. Zhu CH HS XL ZG FJ
Xinzhong Wang BL Yu Zhang JH Jing Wang HZ YW XF GZ Jian Wang
XZ MHS HD Jun Wang Xiaoning Wang.
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Human Exome Capture Chip Applied to Other Primates
PLoS ONE | www.plosone.org 12 July 2012 | Volume 7 | Issue 7 | e40637