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A Reverse Genetic Strategy and Genetic Screen using SNP Genotyping to Reduce Lamb Mortality in Dairy Sheep

Enhancing Animal Breeding Programs

PACE genotyping (PCR Allele Competitive Extension) has become a cornerstone technology in animal genomics, offering precise, rapid, and cost-effective solutions for genotyping. It is widely used to identify and select animals with desirable traits by accurately tracking Single Nucleotide Polymorphisms (SNPs) and insertions/deletions (Indels) associated with essential traits such as disease resistance, productivity, and reproductive performance. This capability allows breeders to make informed decisions when selecting breeding pairs, accelerating the development of improved animal breeds, and enhancing overall breeding programs (you can read a previous blog on this here) .

Recent Advances in Genomic Selection for Livestock Improvement

Over the past decade, genetic selection has significantly improved production traits in livestock. The introduction of new technological tools has further advanced genomic selection, enabling unprecedented genetic progress. High-density SNP chips, improved reference genomes, and detailed gene annotations have facilitated the fine mapping of genomic regions and the identification of causal variants linked to production traits. While these advancements have been successful, they have also been accompanied by challenges, such as a decline in fertility observed in dairy cattle.

Genetic studies have shown that Mendelian monogenic disorders contribute to fertility issues. Although environmental factors play a significant role in reproductive performance, genetic interventions have proven effective in correcting trends and improving fertility, even though its heritability is relatively low (below 0.05).

Classical Forward Genetic Screen

One common method for identifying recessive deleterious variants is the classical forward genetic screen. This approach relies on case-control analysis and genome-wide association studies (GWAS) to distinguish affected from unaffected animals based on distinctive phenotypes and biological samples. Homozygosity mapping is then used to identify homozygous regions in affected animals, which are likely to harbor the causal variant. These findings are often confirmed using whole-genome sequencing (WGS) data. However, this method faces limitations when biological samples and descriptive phenotypes are not available.

Reverse Genetic Approach

To address these limitations, a reverse genetic strategy has also been developed (see reverences below). This approach uses large genomic selection datasets to detect haplotypes with a deficit in homozygous animals, indicating significant deviations from Hardy-Weinberg equilibrium. Originally designed to detect embryonic lethal variants, this method can also fine-map deleterious variants that cause neonatal or juvenile lethality and morphological disorders.

Case Study: Reducing Lamb Mortality in Dairy Sheep

Lamb Mortality Challenges

Lamb mortality poses significant challenges to agriculture, impacting animal health, welfare, and economic outcomes. Early lamb losses, particularly in the first month of life, are often due to a combination of environmental and genetic factors. Recessive deleterious genomic variants, when inherited from both parents, can be lethal, leading to high mortality rates in sheep populations.

INRAE’s Research on Sheep Genomics

The Small Ruminants Genomics team at INRAE, France, is dedicated to identifying genes and their variants underlying traits in sheep and goats and understanding the molecular mechanisms of these phenotypes. The team has developed a comprehensive workflow for identifying and validating genetic variants associated with lamb mortality, specifically focusing on the Manech Tête Rousse and Lacaune dairy sheep breeds.

Identifying Lethal Variants

Through whole-genome sequencing and high-density variant arrays, the INRAE team has been able to fine-map genomic regions and identify causal variants associated with lamb mortality. They have discovered lethal variants in three key genes—MMUT, CCDC65, and SLC33A1—and are actively searching for more. The segregation of these variants is studied across multiple sheep breeds, and their lethal effects are validated through controlled mating trials between heterozygous carriers. Read their case study here.

Implementing PACE Genotyping

To facilitate this research, the team has developed genetic screens using 3CR Bioscience’s PACE® genotyping technology. This technology allows for the quick and cost-effective identification of lethal variants within breeding populations. By screening rams and ewes, breeders can identify individuals carrying potentially lethal variants, thereby controlling mating practices to reduce the occurrence of lethal crosses. This proactive approach improves animal health, welfare, and overall lamb viability.

Broader Applications in Sheep Breeding

Beyond reducing lamb mortality, the workflow and PACE genotyping technology are employed to identify and screen genes affecting other sheep morphological traits, such as horns and coat color. This comprehensive approach ensures that breeders can maintain desired traits while mitigating the risk of genetic disorders. Read a lot more about 3CR Bio’s genotyping technology in action in our Applications sections.

Conclusion

PACE genotyping is a transformative tool in animal genomics, enhancing breeding programs, conservation efforts, and veterinary diagnostics. Its precise, rapid, and cost-effective genotyping capabilities enable breeders to improve animal health, productivity, and welfare. The case study on reducing lamb mortality in dairy sheep exemplifies the significant impact of this technology on animal breeding and highlights the potential for further advancements in the field.

References:

  • Ben Braiek et al. (2024) “Searching for homozygous haplotype deficiency in Manech Tête Rousse dairy sheep revealed a nonsense variant in the MMUT gene affecting newborn lamb viability.” Genetics Selection Evolution.
    • Ben Braiek et al. (2023) “A single base pair duplication in SLC33A1 gene causes fetal losses and neonatal lethality in Manech Tete Rousse dairy sheep.” bioRxiv.
    • Ben Braiek et al. (2022) “A Nonsense Variant in CCDC65 Gene Causes Respiratory Failure Associated with Increased Lamb Mortality in French Lacaune Dairy Sheep.” Genes.
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    Steve Asquith Managing Director
    Steve began his career in the Genetics Division of GlaxoSmithKline, as part of the team establishing GSK’s high-throughput core genotyping laboratory. Steve joined KBioscience when it was first founded in 2002 and was a key driver in taking the company from a small start-up to a multi-national service laboratory, quickly growing the company’s revenue to over $7.5M p.a. Following the acquisition of Kbioscience by LGC in 2011, Steve was appointed Global Director of Operations for LGC Genomics, responsible for over 100 staff in Europe and N. America, successfully elevating the genotyping products and service business. Steve held a crucial leadership role until he left in 2016. In 2017 Steve joined forces with John Holme to create 3CR Bioscience, a new company with a mission to deliver outstanding, customer-focused genotyping products with innovation and affordability at its core.
    Dr. John Holme Technical Director
    John joined KBioscience shortly after it was founded, in 2003, and became Head of Technical Development, building the company’s genotyping and DNA extraction product portfolio and service delivery until 2011 when it was acquired by LGC. Post-acquisition, John was appointed Head of Technical Group for LGC Genomics, in charge of all Research & Development and Technical Support activities for the company. In this role John continued to build on the high-quality products and services provided to the companies growing customer base. During the 19 years John has worked in commercial R&D, he has co-invented numerous highly successful products including PACE®, ProbeSure, KASP™, KlearKall, KlearGene, KlearAmp and KlearTaq™, creating breakthrough offerings in genotyping and extraction and generating huge revenues for the companies he has worked in. In 2017, he joined forces with Steve Asquith and started 3CR Bioscience. John is dedicated to developing outstanding, innovative genotyping products and providing the very best technical support to customers globally.
    Dr. Nisha JainOperations Director
    Nisha has been innovating since the start of her career at Geneform Technologies developing Iso-thermal Genotyping Technologies. Nisha joined KBioscience in 2008, as Senior R&D Scientist and key account Technical Support Scientist, developing KASP and Klearkall performance and coinventing two further versions of KASP. Nisha has more than 15 years’ experience working in molecular biology and genotyping technologies, with extensive experience in the areas of R&D, Quality Assurance and Customer Technical Support. She has technically assisted many giants of the industry with their protocol development and troubleshooting and continues to deliver high-quality support and guidance. In 2018, Nisha joined 3CR Bioscience as Operations Director where she continues to develop PACE and ProbeSure for an increasing range of applications, and to grow 3CR Bioscience’s new product pipeline. Nisha is dedicated to developing outstanding, innovative genotyping products and providing the very best technical support to customers globally.
    Nazma Saffin General Manager
    For 20 years Nazma Saffin has worked and gained extensive expertise within the genotyping sector. Working at Kbioscience and then LGC, she has held operational leadership posts responsible for manufacturing and laboratory services. With experience of ISO 9001 implementation, production scale up and LEAN operations, Nazma has successfully led highly profitable production departments. Joining 3CR Bioscience in 2022, Nazma is committed to delivering operational excellence.
    Jon Curtis Non-Executive Chair
    After 8 years in The Royal Air Force, Jon moved to the Imperial Cancer Research Fund where he pioneered the use of ultra high-throughput genomic automation, capable of 46,000 PCRs per hour. In the 1990’s Jon joined GlaxoSmithKline, implementing a high-throughput genomics platform into their drug discovery pipeline. Whilst there he also developed acoustic mixing into compound management, becoming the gold standard across pharma. Jon developed the world’s first commercially viable 1536-well PCR plates, automated thermal & laser plate-sealer, plus automated liquid-handling & tip washing tools to reduce waste and costs. In 2002 Jon co-founded KBioscience with Phil Robinson, utilising ultra high-throughput PCR instrumentation & a suite of automation tools to create the company’s SNPline robotic platform, with a capacity of 250,000 PCRs/day. The business was underpinned by their ground-breaking patented genotyping chemistry, KASP™, which has over 10,000 scientific papers to date. In November 2022 Jon joined 3CR Bioscience acting as an advisor bringing his commercial and scientific experience to the company.