Genotyping is performed using either of two technologies, Illumina BeadArray for SNPs or ABI capillary electrophoretic sizing of fluorescently tagged PCR products encompassing STRPs or other genomic regions of interest. The Core adopted ABI technology to find indels generated initially by zinc finger nuclease (ZFN) technology, and recently by CRISPR/Cas mutagenesis approach. The Core processed over the last five years 134,000 PCR fragments for zebrafish mutagenesis projects for multiple investigators and the Zebrafish Core. Currently, services include human and mouse SNP genotyping, mouse, zebrafish and human primer testing and (CRISPR/Cas) mutation screening, mouse speed congenics, human and mouse STRPs, MLPA for deletions, and fragment analysis for a variety of other applications. Both Illumina and ABI technologies are widely used by a large number of NHGRI investigators. This year, in addition to two programs (NISC and UDP), a total of 20 investigators representing eight branches used the Core genotyping services. Over the past five years (FY2012-FY2016), there has been a steady increase in the number of DNA samples processed (9,895, 17,608, 40,350, 42,466 and 48,452), and the number for FY2017 is 34,297. There is a continued increase in the number of samples processed for SNP genotyping but a decrease in the number of zebrafish DNA samples processed for mutation screening Human SNP genotyping was carried out on ten different BeadArray types using Illumina Infinium technology. We processed, this year, 8,743 human DNA samples including 384 samples processed using human methylation arrays, and generated 10 billion genotypes. Last year (FY2016), we processed 7,356 human DNA samples. The genotyping data generated this year are used for studies related to Proteus syndrome, Diamond-Blackfan Anemia, Fanconi anemia, Mobius syndrome, cancer, Inherited bone marrow failure syndromes, Smith Magenis syndrome, Cardiovascular diseases, Scleroderma, Gaucher Disease/Parkinson Disease, Congenital Heart Disease study, Turner syndrome, ADHD and Twins Study, among others. The data were analyzed for identity by descent, copy number variation, deletion intervals, methylation status, parent-of-origin of deletions, mosaicism, and to generate haplotypes for discovering variants from sequence data. In addition to numerous small projects, we do have some large SNP projects. Over the last five years (FY2013-FY2017), the number of samples we processed for NISC were 1,536, 2,256, 4,032, 2,256, and 2,712. We understand that the genotyping samples processed for NISC belong to multiple investigators from other institutes, indicating the Genomics Core serves a larger scientific community than just NHGRI. Further, a large SNP genotyping project for Cardiovascular diseases started two years ago was extended to another 1,448 DNA samples this year, adding to a total of over 5,000 DNA samples. Over the last two years, 2,944 DNA samples were processed for the scleroderma studies from another lab. Samples run on SNP and methylation arrays (8,743 samples) represent about 25% of the total 34,297 DNA samples processed by the Core this year, which is substantially higher than the 15% observed last year. The remaining samples (25,554) were processed using ABI technology, which has the capability to separate fluorescently labeled PCR products at single-base resolution. An efficient screening strategy for identification of germline transmitting founder fish and the size of insertion/deletion mutations generated by ZFNs and TALENs, started as a modest effort in 2010, has had a huge surge due to the introduction of CRISPR/Cas technology for targeted mutagenesis. The Genomics Core has processed a large number of zebrafish DNA samples (14,619, 35,539, 28,986, 34,714, and 20,198) respectively for the FY2013-2017. Though the number of zebrafish samples decreased this year, the CRISPR technology is being extended to an increasing number of mouse mutagenesis samples. The Core has been assisting investigators with data analysis and access to software/tools, such as GoldenHelix, Nexus, and GenomeStudio. The yearly renewal expense of Nexus and GoldenHelix limits their availability to only one computer in the Core. The Core helps researchers to take advantage of learning and using these tools, and also helps with the handling, collection, evaluation, and processing of SNP and other data sets. The Core has provided significant data analysis support over the past four years (FY2013-FY2016) (Table 2a, b), as well as this year (FY2017) (Table 2c). The services are related to copy number variation, linkage disequilibrium analysis, population stratification, and association studies. Analyses for detecting deletions, duplications, loss of heterozygosity, and regions portraying signs of chromosomal mosaicism in DNA samples from patients diagnosed with Fanconi anemia and head and neck cancer were also performed. Other studies include changes associated with the processing of iPS, Acute myeloid leukemia, Smith-Magenis syndrome, Febrile Infection-Related Epilepsy Syndrome, population stratification and variant frequency ranking of the African-American Ancestral SNPs, and eye diseases Coloboma and Leber Congenital Amaurosis (LCA), ADHD, Congenital Heart Disease, Holoprosencephaly, Turner syndrome, and Fanconi anemia. In addition to performing analysis, the core also helps with troubleshooting or problem solving any issues investigators may have in handling their data. This service is of huge value to investigators with small projects, as are most users of the Core, who do not have the required tools or expertise for the analysis of large data sets. Over the last two years, we added six new genotyping panels: HumanOmni2.5Exome, African Diaspora, MEGA Consortium, Methylation EPIC, CVD_Custom, and HumanOmni5Exome. These new array types involved a new LCG technology that created a few initial hurdles that needed to be resolved. We also started processing custom SNP chips in the FY2017: MEGA, CVD, and OmniExpressExome. Anticipated growth of sample numbers will continue next year. Fragment analysis is being used increasingly for zebrafish and mouse CRISPR based mutagenesis and thus anticipated use of the Genomics Core for mutation screening will continue to be large. SNP genotyping will continue to increase in the coming years. NISC expects a large increase year after year. Another large sample size project was started in FY2017, and processing samples for this project will continue in the coming year.
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