Rhesus macaque 96 SNP parentage assay:
Genotyping for parentage validation is performed using a 96 SNP assay, performed on a Fluidigm Biomark HD platform. This SNP assay was developed in partnership with the NPRC Genetics and Genomics Working Group Consortium to enable consistent genotyping and comparison of rhesus macaques across all NPRCs. Quality Control: two replicate samples are included with each assay to evaluate the reproducibility of genotype calls.
Rhesus macaque SNP ancestry assay:
This SNP assay includes 122 Ancestral Informative Markers (AIMs) selected for high information content and broad genomic representation. The genotyping is carried out using the ABI QuantStudio 12K Flex System. The genotypes are then compared with 74 animals of known origin (35 Indian rhesus, 35 Chinese rhesus, and 4 hybrid animals of mixed Indian/Chinese origin). STRUCTURE analysis (Pritchard et al., 2000) is used to calculate a statistical probability of assignment into the two populations, with the 90% probability interval reported. For quality control: two previously genotyped DNA samples are included to evaluate consistency of genotype and ancestry calls. Ancestry determination is not made on DNAs with more than 10% missing genotype information.
For projects and assays not requiring the throughput of the NovaSeq 6000 operated by the OHSU Massively Parallel Sequencing Shared Resource, the Primate Genetics Core has options for NGS (Illumina MiSeq) and capillary sequencing (ABI 3730xl). We specialize in working with researchers to optimize assays to meet research needs in a timely fashion.
MHC expressed allele assay:
Macaque MHC class I genotyping is performed using the RNA-based expressed allele assays, as described by Wiseman et al., 2009. Targeted polymorphic MHC Class I regions are amplified, multiplexed and sequenced on an Illumina MiSeq sequencer. The resulting reads are compared against all defined MHC alleles to identify alleles and haplotypes. Alleles with a minimum of 0.2% sequence representation are reported. Finalized MHC alleles and haplotypes are reported directly or uploaded into the PRIMe database. Quality Control: A previously analyzed RNA sample is included in each round of library construction to ensure the reproducibility of results.
Custom genotyping assays:
The design of assays to distinguish macaque Single Nucleotide Variants (SNVs) or Variable Number Tandem Repeats (VNTRs) are available for investigators by request. Genotyping based on previously established assay designs (CLN7 c.769delA, 5-HTTLPR, MAOA-LPR, CRH C-2610T, TPH2 A2051C, Mamu-A*01, Mamu-B*08, Mamu-B*17) are also available. DNA samples with known genotypes are included to assure the reproducibility of assay results.
ONPRC NHP DNA Bank:
The ONPRC DNA Bank contains whole blood and liver samples collected on an ongoing basis from all rhesus and Japanese macaques to support internal and external research programs. DNA is extracted as needed from buffy coat, whole blood, or liver samples. Currently, the DNA Bank includes over 50,000 archived samples from more than 19,000 NHPs, including 11 different species.
Other services and self-service instumentation:
Most specialized and general equipment maintained and operated by the core is also available for use by individual investigators. Training, supervision, and necessary reagents is supplied. Available equipment includes real-time PCR machines, automated nucleic acid extraction robots, DNA/RNA QC platforms, plate readers, cell counters, and gel imagers. Please inquire about other services or requests.
Nucleic acid extraction and quality control:
The Primate Genetics Core has extensive experience extracting DNA and RNA from a variety of tissue sources using standard protocols and automated systems. Quality and quantity of isolated samples is assayed with ThermoFisher Qubit Fluorometer, Nanodrop Spectrophotometer and Agilent Bioanalyzer.