More recent methods have been developed, which use WGS as an initial step but such methods typically require additional resources or sequencing a large number of animals. The second stage used a range of “fine-mapping” methods to search within the highlighted region for the site of the new variant, many of which resulted in whole genome sequencing (WGS) of a few cases and controls. First, the use of a suitable single-nucleotide polymorphism (SNP) chip in a case/control study to locate the region containing the new variant combined with a method searching for long runs of homozygosity (ROH), the more traditional chi-squared method being shown to be inadequate. This should speed-up and simplify the management of new genetic diseases to a single-step process.Ī review of 34 articles detailing work to map 38 novel autosomal recessive genetic conditions to their position on the genome ( Pollott, 2018) suggested that finding the site of such a condition required the use of at least a two-stage methodology.
In many instances it may also be confirmed with variant prediction scoring. The genomic site of novel autosomal recessive Mendelian genetic diseases can be located using fewer than 20 animals combined with two bioinformatic methods, autozygosity-by-difference, and genotype criteria. Fewer than 20 animals need to be sequenced when using the GCR and ABD methods together. Locating GCR sites was best performed using two carriers to every case, and the carriers should be distantly related to the cases, within the breed concerned. Investigation of the GCR method found that sites meeting the GCR were not evenly spread across the genome but concentrated in regions of long runs of homozygosity. Both methods confirmed the candidate site.
It was verified on an independent sample of animals from the breed using genotyping by polymerase chain reaction at the candidate site and autozygosity-by-difference using SNP-chips. This site was a splice acceptor variant located in the glucokinase gene ( GCK). From more than nine million variants in the VCF file, only one site was identified by all four methods (Chr4: g.77173487A>T (ARS-UCD1.2 (GCF_002263795.1)). Four methods were used to interrogate the variant call format (VCF) data file of these nine animals, they are genotype criteria (GCR), autozygosity-by-difference (ABD), variant prediction scoring, and registered SNP information.
Using whole genome sequencing of only three cases and six carriers, the site of a novel variant causing perinatal mortality in Irish moiled calves was located.
#Allocating more memory into clc genomics workbench full#
This study explores bioinformatic methods that can be used to identify the causative variants of recessive genetic disorders with full penetrance with just nine whole genome-sequenced animals to simplify and expedite the process to a one-step procedure. This region is then explored using fine-mapping methods to locate the actual site of the new variant. Until recently, finding the causal genomic site of a new autosomal recessive genetic disease has required a two-stage approach using single-nucleotide polymorphism (SNP) chip genotyping to locate the region containing the new variant. To manage them effectively, some methods need to be devised that are quick and accurate. New Mendelian genetic conditions, which adversely affect livestock, arise all the time. 4The Roslin Institute, Midlothian, United Kingdom.3Department of Pathobiology and Population Sciences, The Royal Veterinary College, Hatfield, United Kingdom.2Comparative Neuromuscular Diseases Laboratory, Department of Clinical Sciences and Services, Royal Veterinary College, London, United Kingdom.1Department of Pathobiology and Population Sciences, The Royal Veterinary College, London, United Kingdom.Piercy 2, Claire Massey 2, Mazdak Salavati 3,4, Zhangrui Cheng 3 and D.
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