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Building scientific studies

Every gain in knowledge in animal genetics rests on careful collaboration: between scientists, breeding organisations and – above all – the individual breeders. The laboratory contributes the methods and the computing power. Whether a project succeeds or fails, however, is decided long before that: in how the animals are described, and in what material is available from them.

This page sets out how a robust study comes about – from precise phenotyping through the form of organisation and the sampling strategy to the association study itself.

Precise phenotypes – identifying cases and controls

An association study compares animals that show a trait with animals that do not. Everything the statistics can achieve later is decided by how reliably those two groups are formed. An animal placed in the wrong group does not simply add noise – it removes power. The phenotype, not the genotype, is the part of a study that breeders and breeding organisations control.

Criterion 1

Define the case in writing

Before the first sample is drawn, it must be written down what counts as affected: which finding, established by which method, at what age. A diagnosis that rests on an impression cannot be reproduced by a second examiner – and therefore cannot be analysed.

Criterion 2

A control is not simply an animal without a report

Controls must be examined by the same protocol as the cases. For traits that appear late, the control must have reached an age at which the trait would have become visible. A young, unexamined animal is not a control – it is an unknown.

Criterion 3

Keep the grade, do not reduce it to yes/no

Where a graded assessment already exists – the FCI grades A to E for hip dysplasia, for example – it should be carried into the analysis as a grade. Collapsing it into affected/unaffected throws away information that has already been paid for.

Bar chart: with a clean case definition 486 animals are needed, with one per cent of healthy animals wrongly classified as cases 921 animals are needed, an increase of 89 per cent.
Sample size required for the same statistical power, after Edwards et al. (2005).

Checklist for a usable phenotype

  • A written case definition, agreed before the first sample.
  • A standardised examination: who examines, by which method, at what minimum age.
  • The same protocol for cases and controls.
  • Date of examination, age of the animal and the examiner documented with the finding.
  • Graded findings carried over as grades, not reduced to a binary.
  • Stated exclusion criteria – trauma, competing diagnoses, incomplete examination.
  • Unambiguous animal identity, so that finding and sample belong to the same animal beyond doubt.

Standardised assessment also makes data comparable across borders. Wang et al. (2021) combined 1,357 German Shepherd dogs from the United Kingdom, Sweden and Finland into one hip dysplasia study although two different scoring schemes were in use. The same work provides the empirical counterpart to the argument above: using the graded FCI classification produced more significant and suggestive SNPs than the binary case/control split on the very same animals.

Sources

  • Edwards BJ, Haynes C, Levenstien MA, Finch SJ, Gordon D (2005): Power and sample size calculations in the presence of phenotype errors for case/control genetic association studies. BMC Genetics 6:18. doi.org/10.1186/1471-2156-6-18
  • Wang S, Strandberg E, Arvelius P, Clements DN, Wiener P, Friedrich J (2021): Genome-wide association studies for canine hip dysplasia in single and multiple populations – implications and potential novel risk loci. BMC Genomics 22:636. doi.org/10.1186/s12864-021-07945-z

The right organisation – where observations become data

One breeder's observation is the beginning of every study – and at the same time the most fleeting part of it. For it to become usable data, something has to bring findings and animals together over years, secure the identity of the animals and govern who may see what. That is exactly what DNA programmes and the Animal Trust Center (ATC) are built for.

Role 1

The individual breeder

They see what nobody else sees: the puppy that develops differently, the bitch that fails to conceive a second time, the foal with the conspicuous gait. What matters is to report the unremarkable animals as well. Passing on only the problem cases delivers a study without controls – and therefore no study at all.

Role 2

The breeding organisation

The organisation is the only place with a complete view of a population: studbook, pedigrees, litter registrations and mandatory examinations all come together there. It determines which examination is compulsory for which animal – and in doing so creates a uniform phenotype in the first place.

Role 3

DNA programmes

A breeding organisation stores its own DNA programme in the ATC: which tests apply to which animals, and which findings are shared within the organisation. Many individual orders thereby become one continuous, uniform data collection.

Role 4

Animal Trust Center

In the ATC, animals and findings are held in a digital online animal record, every finding carrying a secure electronic signature. The owner decides what stays private and what is shared with the organisation – a study is therefore never built behind the breeders' backs, but with their explicit release. See the account types

Sample material – two strategies

Whether a question can be answered later depends on whether the material for it exists. There are two fundamentally different routes to sampling. Both are legitimate; they do not differ in quality but in how many questions they keep open.

Two populations side by side: under project-based sampling only the affected animals and their controls are sampled, under continuous archiving all breeding animals are; when a new question arises, the first has no material available and the second has it immediately.
The same population, two sampling strategies – and what happens when a new question comes up two years later.
Strategy A

Sample for the project

Only those animals are sampled that have been identified as affected by a particular trait, together with their matching controls. The collection begins with the question and ends with it.

Advantage

  • Costs stay limited to a few animals and can be calculated in advance.
  • The organisational effort is manageable and has a defined end.
  • For a clearly outlined question that has already been formulated, it is the fastest route.

Disadvantage

  • No further use: every new question requires a new collection.
  • Nothing can be made up retrospectively – animals have been sold, have died or can no longer be sampled.
  • The control group is tailored to the original question and is rarely reusable.
Strategy B

Archive continuously

A sample is taken and stored from every breeding animal – whether or not a project is currently running. The archive grows with the population and is not committed to any one trait.

Advantage

  • What will be looked for is not fixed in advance: every later question finds material waiting.
  • Retrospective analyses become possible, because earlier year groups are included.
  • Cases and controls can be selected to fit afterwards instead of being defined up front.
  • The population is represented completely – including the unremarkable animals.

Disadvantage

  • Higher costs, because every breeding animal is sampled and not only those needed.
  • Ongoing organisational effort and permanent storage.
  • Part of the material will never be used – and which part cannot be known beforehand.

Source

  • Groeneveld LF, Gregusson S, Guldbrandtsen B, Hiemstra SJ, Hveem K, Kantanen J, Lohi H, Stroemstedt L, Berg P (2016): Domesticated Animal Biobanking: Land of Opportunity. PLOS Biology 14(7):e1002523. doi.org/10.1371/journal.pbio.1002523

Association studies – once enough informative samples exist

An association study compares marker genotypes between cases and controls across the whole genome and asks where the two groups differ systematically. It becomes possible at the moment when enough informative samples have accumulated. How many that is depends on the trait, not on the budget – and the difference between a monogenic and a complex trait is not a matter of degree but of orders of magnitude.

~20

Animals for a mendelian trait

Karlsson et al. (2007) mapped mendelian traits within a breed using an array of roughly 27,000 SNPs and about 20 dogs, reaching regions below 1 Mb. Fine mapping across two breeds then narrowed the S locus to approximately 100 kb within MITF. The strategy that follows from this is: map within the breed, fine-map across breeds.

4,200+

Animals for complex traits

Hayward et al. (2016) genotyped more than 4,200 dogs at 180,000 markers and found significant loci for hip dysplasia, elbow dysplasia, idiopathic epilepsy, lymphoma, mast cell tumour and granulomatous colitis. Anyone expecting an answer on hip dysplasia from 30 samples is working three orders of magnitude below what the trait requires.

100×

Why dogs need fewer animals

Sutter et al. (2004) measured linkage disequilibrium within breeds reaching from 400–700 kb in Golden and Labrador Retrievers to 3–3.2 Mb in Bernese Mountain Dogs and Pekingese – up to a hundred times further than in humans, so correspondingly fewer markers are needed. The range is breed-specific, however: between those breeds it differs roughly fivefold.

From the array to the sequence

SNP arrays are sufficient for finding associations within a breed. Across several breeds, and in mixed-breed animals, linkage disequilibrium decays between the markers and the array runs out of power. Two developments have changed this.

Imputation against a sequence panel

Hayward et al. (2019) imputed array genotypes against 365 whole-genome-sequenced dogs and wolves and raised the number of usable markers from 173,000 to 24.0 million, at an overall accuracy of 88.4 per cent (purebred dogs 89.7 per cent, village dogs 84.2 per cent). The gain was not theoretical: a new significant locus for cranial cruciate ligament disease appeared on CFA20.

Reference panels of a different size

The Dog10K consortium sequenced 1,987 canids at twentyfold coverage and described more than 48 million variants (Meadows et al. 2023). In the horse the resources are markedly weaker: Reich et al. (2022) reached an imputation accuracy of only 0.66 from a 60K array against 317 horse genomes, but around 0.90 from a 670K array. For horses the shortcut via a sparse array does not carry.

Sources

  • Sutter NB, Eberle MA, Parker HG, Pullar BJ, Kirkness EF, Kruglyak L, Ostrander EA (2004): Extensive and breed-specific linkage disequilibrium in Canis familiaris. Genome Research 14(12):2388–2396. doi.org/10.1101/gr.3147604
  • Karlsson EK, Baranowska I, Wade CM, Salmon Hillbertz NHC, Zody MC, Anderson N, Biagi TM, Patterson N, Rosengren Pielberg G, Kulbokas EJ, Comstock KE, Keller ET, Mesirov JP, von Euler H, Kämpe O, Hedhammar Å, Lander ES, Andersson G, Andersson L, Lindblad-Toh K (2007): Efficient mapping of mendelian traits in dogs through genome-wide association. Nature Genetics 39(11):1321–1328. doi.org/10.1038/ng.2007.10
  • Hayward JJ, Castelhano MG, Oliveira KC, Corey E, Balkman C, Baxter TL, Casal ML, Center SA, Fang M, Garrison SJ, Kalla SE, Korniliev P, Kotlikoff MI, Moise NS, Shannon LM, Simpson KW, Sutter NB, Todhunter RJ, Boyko AR (2016): Complex disease and phenotype mapping in the domestic dog. Nature Communications 7:10460. doi.org/10.1038/ncomms10460
  • Hayward JJ, White ME, Boyle M, Shannon LM, Casal ML, Castelhano MG, Center SA, Meyers-Wallen VN, Simpson KW, Sutter NB, Todhunter RJ, Boyko AR (2019): Imputation of canine genotype array data using 365 whole-genome sequences improves power of genome-wide association studies. PLOS Genetics 15(9):e1008003. doi.org/10.1371/journal.pgen.1008003
  • Meadows JRS, Kidd JM, Wang G-D, Parker HG, Schall PZ, Bianchi M, Christmas MJ, Bougiouri K, Buckley RM, Hitte C et al. (2023): Genome sequencing of 2000 canids by the Dog10K consortium advances the understanding of demography, genome function and architecture. Genome Biology 24:187. doi.org/10.1186/s13059-023-03023-7
  • Reich P, Falker-Gieske C, Pook T, Tetens J (2022): Development and validation of a horse reference panel for genotype imputation. Genetics Selection Evolution 54:49. doi.org/10.1186/s12711-022-00740-8

Publications

Association studies are carried out together with university research groups, and the result belongs in the scientific literature. The following selection shows papers in which Generatio scientists appear as authors or co-authors – chosen for their relevance to the methods described on this page.

Eberhard Manz

Founder and Managing Director, Generatio GmbH – Center for Animal Genetics

  • Tan K, Adeniyi OO, Letko A, Manz E, Wagner HW, Zanolari P, More M, Gutierrez-Reynoso GA, Drögemüller C, Lühken G (2026): Genome-wide SNP-based genomic diversity and population structure analysis in alpaca populations from Europe and Peru. G3 (Bethesda) 16(9):jkag157. doi.org/10.1093/g3journal/jkag157
  • Rietmann SJ, Lange A, Soto S, Thom N, Manz E, Jagannathan V, Mayer U, Leeb T (2024): KRT5 in-frame deletion in a family of German Shepherd dogs with split paw pad disease resembling localized epidermolysis bullosa simplex in human patients. Animal Genetics 55(4):692–696. doi.org/10.1111/age.13444
  • Tan K, Adeniyi OO, Letko A, Rudd Garces G, Manz E, Wagner H, Zanolari P, Drögemüller C, Lühken G (2024): Identification of genomic regions associated with differences in fleece type in Huacaya and Suri alpacas (Vicugna pacos). Animal Genetics 55(1):163–167. doi.org/10.1111/age.13377
  • Tan K, Roy M, Manz E, Wagner H, Zanolari P, Drögemüller C, Lühken G (2022): The KIT:c.376G>A variant in German and Swiss alpacas (Vicugna pacos) with different coat colors. Animal Genetics 53(5):718–720. doi.org/10.1111/age.13231
  • Manz E, Tellhelm B, Krawczak M (2017): Prospective evaluation of a patented DNA test for canine hip dysplasia (CHD). PLOS ONE 12(8):e0182093. doi.org/10.1371/journal.pone.0182093
  • Gerber M, Fischer A, Jagannathan V, Drögemüller M, Drögemüller C, Schmidt MJ, Bernardino F, Manz E, Matiasek K, Rentmeister K, Leeb T (2015): A deletion in the VLDLR gene in Eurasier dogs with cerebellar hypoplasia resembling a Dandy-Walker-like malformation (DWLM). PLOS ONE 10(2):e0108917. doi.org/10.1371/journal.pone.0108917

Earlier work at Hannover Medical School dealt with Y-chromosomal gene families in humans and cattle and with DNA fingerprinting for parentage verification – among others Nürnberg P et al. (1998), Paternity assessment in rhesus macaques (Macaca mulatta): multilocus DNA fingerprinting. American Journal of Primatology 44(1):1–18.

Arnav Mehrotra

Bioinformatics and quantitative genetics, Generatio GmbH – Center for Animal Genetics

  • Gorssen W, Kadri NK, Khayatzadeh N, Leonard AS, He Q, Mehrotra A, Neuenschwander S, Pausch H (2026): Genetic analysis of iris pigmentation in Swiss pig breeds identifies a missense KITLG variant as a potential causal factor for pale and heterochromatic irises. Genetics Selection Evolution 58(1):22. doi.org/10.1186/s12711-026-01040-1
  • Nosková A, Mehrotra A, Kadri NK, Lloret-Villas A, Neuenschwander S, Hofer A, Pausch H (2023): Comparison of two multi-trait association testing methods and sequence-based fine mapping of six additive QTL in Swiss Large White pigs. BMC Genomics 24:192. doi.org/10.1186/s12864-023-09295-4
  • Mehrotra A, Bhushan B, Kumar A, Panigrahi M et al. (2022): A 1.6 Mb region on SSC2 is associated with antibody response to classical swine fever vaccination in a mixed pig population. Animal Biotechnology 33(6):1128–1133. doi.org/10.1080/10495398.2021.1873145
  • Mehrotra A, Bhushan B, Karthikeyan A, Singh A, Panda S, Bhati M, Panigrahi M, Dutt T, Mishra BP, Pausch H, Kumar A (2021): Genome-wide SNP data unravel the ancestry and signatures of divergent selection in Ghurrah pigs of India. Livestock Science 250:104587. doi.org/10.1016/j.livsci.2021.104587
  • Singh A, Kumar A, Mehrotra A et al. (2021): Estimation of linkage disequilibrium levels and allele frequency distribution in crossbred Vrindavani cattle using 50K SNP data. PLOS ONE 16(11):e0259572. doi.org/10.1371/journal.pone.0259572

Before joining Generatio, Arnav Mehrotra worked in the animal genomics group at ETH Zurich and at the Indian Veterinary Research Institute. His focus – association studies, signatures of selection, sequence-based fine mapping – is exactly the set of methods described on this page.

A selection, not a complete bibliography. Search performed in Europe PMC and Crossref, September 2026. Each paper remains permanently retrievable through the DOI given.