Primate Genetics Core
MOLECULAR GENETIC SERVICES
The Molecular Genetics Services group provides research expertise to develop and implement high-throughput macaque genotype analysis to meet both research and colony management needs. Current genotyping services include: rhesus macaque 96 SNP parentage analysis, rhesus macaque 122 SNP ancestry analysis, MHC Class I allele analysis, custom genotype assay development and analysis, and the construction of genotyping-by-sequencing libraries. In addition, this service group also manages an extensive ONPRC non-human primate DNA Bank, which supports both research and colony management needs. DNA or RNA extraction services are also available upon request.
Betsy Ferguson, Ph.D., Director
Rita Cervera-Juanes, Ph.D., Research Assistant Professor
Gwen Steinbach, B.S., R.N., Research Assistant 2
Quynh Quach, B.S., Research Assistant 2
Samantha Martin, B.S., Research Assistant 2
- 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 QuantStudioTM 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. Quality Control: Two previously genotype DNAs are included to evaluate consistency of genotype and ancestry calls. Ancestry determination is not made on DNAs with more than 10% missing genotype information.
- MHC expressed allele assay: The RNA-based MHC assay used to detect expressed class I alleles in rhesus macaques, 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 uploaded into the PRIMe database. Quality Control: A previously analyzed RNA sample is included in each round of library construction to insure reproducibility of results. Samples producing fewer than 1,000 reads are not reported.
- 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. Quality Control: control gDNAs having known genotypes are included to assure reproducbility 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 22,000 archived samples from more than 6,600 NHPs, including 11 different species.
- NHP DNA and RNA extractions: Nucleic Acid extraction is performed on liver, brain, blood or other tissues provided by investigators. Quality Control: DNA and RNA quality is assessed using an Agilent 2100 Bioanalyzer. Quality standards include RNAs having a minimum RIN score of 7.0 and DNAs having a 260/280 ratio within 1.8-2.0.
COLONY GENETIC ANALYSIS SERVICES
The Colony Genetics Resource Core oversees all population genetic analysis of non-human primates (NHPs) at the ONPRC, in order to characterize the large colony pedigree (~23,000 rhesus macaques since 1963), to monitor colony genetic diversity, and to characterize genetic contributions to disease traits in colony macaques. Analyses based on the ONPRC pedigree are used to support colony management decisions related to the formation of breeding groups, and to inform other management strategies that impact the genetic health and diversity of NHPs at the ONPRC. With phenotype and genotype data, the ONPRC pedigree is also an immensely valuable resource for discovering genetic contributions to diseases that are conserved between macaques and humans.
Amanda Vinson, Ph.D., Director
Elizabeth Slikas, Ph.D., Research Associate
- Parentage Assignment and Other Kinship Analysis: Parentage assignment is evaluated using the 96 SNP parentage genotype data, together with the NPRC Consortium parentage analysis pipeline. We also provide analyses of kinship among specified animals based on the overall colony pedigree, as needed by ONPRC investigators.
- Analysis of Population Genetic Diversity: Population genetic metrics include the estimation of kinship for individuals relative to the rest of the colony, the overall kinship within the colony, and probabilistic measures of allelic diversity for individual animals. These metrics are used to identify animals that are the most genetically valuable to the colony, so that breeding groups can be developed that minimize the loss of genetic diversity from the colony as a whole. To support these efforts, we have developed new software to estimate these metrics, and to aid in developing new breeding groups. These results are provided, along with recommendations for new breeding groups, to the Division of Comparative Medicine for use in colony management decisions.
- Maintenance of Colony Genotype Data: The PRIMe database is the central electronic health records (EHR) system for all NHPs at the ONPRC. PRIMe serves as an accessible repository for all genotype and parentage assignment data. Additionally, the continuously updated pedigree permits current estimates of mean individual kinship relative to the colony, as well as pairwise kinship between any 2 macaques, to be accessed through PRIMe.
Charges for obtaining DNA from the DNA bank is $2/ug DNA for internal investigators. If DNA extraction from a buffy coat or liver sample is required, a $20 fee is charged, with the agreement that 10 ug is delivered to the investigator and the remaining DNA sample is retained by the DNA Bank.