99 Lives Cat Genome Sequencing Initiative

 – a public effort to discover cat genetic variation and to improve feline & human health!

Precise determination of the genome of a species is important for the understanding of disease and for the development of diagnostic and screening tests that will improve disease treatment. The biology of one species can assist the understanding of biology, physiology and genetic disease in another species. The costs of genome sequencing have been drastically reduced and genomes are now developing for most every species. A high quality, long-read genome assembly of Cinnamon, the an Abyssinian cat from the University of Missouri, was produced in 2017. This improved genome has made Precision Medicine feasible for the domestic cat.

The 99 Lives Cat Genome sequencing Initiative will continue to support improvements in the cat genome assembly and apply Precision Medicine to cats:

• Improve coverage of the cat genome using newly developing technologies
• Improve future assemblies of the cat genome by producing data for annotation
• Identify normal and abnormal genetic variation
• Identify causative variants for specific health concerns
• Allow veterinary hospitals to provide individual genome sequencing for cats for state of the art health care (similar to the developing standard of care for humans)

Coordination by the Lyons Feline Genetics Laboratory at MU will ensure:

• Different cats from different breeds and locations are sequenced to maximize variation detection
• Similar technologies are used to facilitate efficient data management, transfer and variant identification
• Data quality is high and consistent with standards similar to human-based analyses
• Data is publicly available so that all members of the research community can benefit in a non-competitive and collaborative manner
• Researchers less familiar with genomics research and large scale bioinformatics will have a resource for data interpretation

A FaceBook and other website pages support the project and help to raise funds from the community including, public, private, university and government funding. Donations and acquired funding for the 99 Lives Project will support:

• Payment to veterinary clinics for collection of a 3 – 6 ml EDTA blood sample and overnight shipment to the Lyons laboratory at MU for DNA isolation
• DNA isolation using organic preps (Phenol:chloroform) by the Lyons Laboratory
• Delivery of DNA to library preparation and sequencing facility
• One library preparation (350 – 550 bp)
• Illumina XTen sequencing: 150 bp paired-end reads
• Variant analysis following GATK Best Practices to the cat Version 9.0 assembly
• Maintenance of a DropBox site for distribution of VCF
• Lyons laboratory maintenance of project websites
• Hard drives for data storage, transfer of data amongst participating laboratories
• Supportive research to improve genome assembly and annotation
• Validation of candidate DNA variants for diseases and traits

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Memo of Understanding (MOU) – 2019

Precise determination of the genome of a species is important to understanding disease and for the development of diagnostic and screening tests that will improve disease treatment. The genome has been sequenced in humans, dogs, horses, cattle, cats, and several other species. Unfortunately, the genomic sequence has only been determined in a single cat. Cinnamon was an Abyssinian cat from the University of Missouri whose genome was sequenced in 2007. Because the genome was mapped in a single animal, and since less robust methodologies were used, we lack the information needed to develop tests that may help in the identification of mutations and treatment of disease.

A research community-based project has developed that will sequence the genome of 99 or more cats in order to:

• Improve coverage of the cat genome
• Improve future assemblies of the cat genome
• Identify normal and abnormal genetic variation
• Identify causative mutations for specific health concerns
• Allow veterinary hospitals to provide individual genome sequencing for cats for state of the art health care (similar to the developing standard of care for humans)

The 99 Lives project is being coordinated by the Lyons Feline Genetics Laboratory at the University of Missouri – Columbia to help ensure that:

• Different cats from different breeds and locations are sequenced to maximize variation
• Similar technologies are used to facilitate efficient data management, transfer and variant calling
• Data quality is high and consistent
• Data is publically available so that all members of the research community can benefit in a non-competitive and collaborative manner
• Researchers less familiar with genomics research and large scale bioinformatics will have a resource for data interpretation
• Follow standards and precedence set by human genetics efforts

All data from the project will become publically available, participants that agree to this MOU will have access to data early and prior to publications.

99 Lives Researcher Participation Expectations

Participants in the project are expected to provide high quality sequence and deep coverage of a unique cat genome as part of the collaboration.

• Illumina HiSeq 100 bp paired end sequencing reads are supported. Sequencing is performed from two PCR-free libraries per cat of 350 bp and 550 bp for at least 15X coverage; 20X coverage for standard libraries*.
• Illumina XTen 150 bp paired end sequencing reads are supported. Sequencing is performed from one PCR-free libraries per cat for at least 20X – 30X coverage.
• Contributors must provide the sequencing details for each genome. *Ajay SS, Parker SC, Abaan HO, Fajardo KV, Margulies EH. Accurate and comprehensive sequencing of personal genomes. Genome Res. 2011 Sep;21(9):1498-505. doi: 10.1101/gr.123638.111. http://genome.cshlp.org/content/21/9/1498.long
• The data needs to be transferred to the Lyons Laboratory at MU for variant calling against cat assembly version 9.0. Variant calling will occur approximately every 6 months.
• Once variants have been called across the entire dataset – participants will be provided a link to obtain the VCF.

Contributors are expected to provide the basic signalment of the cats when possible, such as gender, breed, place of origin, and coat color.

• One of the most significant concerns of a shared resource is the potential for overlapping projects and performing data analyses on contributed genomes that would interfere or “scoop” the research of another contributor. Therefore, the specific phenotype(s) of interest can be withheld from the cat identification information. However, we fully expect a highly cooperative and collaborative effort for participation in the resource.
• Researchers can privately contact a subset of contributors to ask for phenotypes of interest and for collaborative efforts.
• All cats are welcome, including exotic and wild felids.

All contributors to the 99 Lives data resource are expected to be co-authors on the first publication that is produced by each contributor who uses the data resource.

• All funding sources need to be provided for acknowledgements and included in each publication and presentation using the 99 Lives resource data.
• All the cat genome data can be downloaded by other investigators who are interested in other data analyses of the cat genome provided they make a contribution to the resource.
• Cat sequences will be deposited into NCBI or the European Nucleotide Archive < EMBL-EBI

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99Lives Cat Genome Sequencing Initiative Contributors

The project has developed as collaboration between the University of Missouri – Columbia, College of Veterinary Medicine, and the University of California – Davis but is extended to all researchers interested in feline health care. The Feline Genetics Laboratory of Leslie Lyons, PhD, at the University of Missouri is coordinating the project. For details – please to e-mail; lyonsla@missouri.edu or felinegenome@missouri.edu or call 01 573 884 2287.

The Lyons’ Feline Genetics Laboratory will provide periodic updates regarding the project to the contributors and the research community.

The contributors listed below have agreed to the MOU and have submitted or are anticipating the contribution of cat genome sequences to the 99 Lives Cat Genome Sequencing Initiative.

Table 1. 99 Lives Institutions and Researcher Contacts

*Institution at time of genome submission

**Current email address

^†Submission of wild felid genomes