Supplementary Table 1 . Numbers of Dogs and Pedigree Designs for Labrador Retriever-Derived

12/9/18

Supplementary Table 1. Numbers of dogs and pedigree designs for Labrador retriever-derived (drd1) and Samoyed-derived (drd2) Oculoskeletal Dysplasia (OSD) informative canine pedigrees.

Disease / DNA samples sent to Genotyping / Pedigree design
Dogs normal for OSD / OSD affected dogs / OSD obligate heterozygotes (carriers) / Non-dwarf dogs from inter-cross breeding / Intercross breeding (offspring) / Back-cross breeding (offspring)
drd1 / 4 / 30 / 14 / 32 / 5 (50) / 3 (18)
drd2 / 3 / 23 / 16 / 28 / 2 (42) / 3 (21)

Supplementary Table 2. Numbers of purebred dogs tested for the presence of COL9A3 and COL9A2 OSD-associated mutations. (A) Labrador retrievers tested for COL9A3 mutation, their phenotype and results. (B) Samoyeds tested for COL9A2 mutation, their phenotype and results. (C) Dogs from breeds not known to segregate OSD tested homozygous normal for COL9A3 and/or COL9A2 mutations. (D) Dogs from breeds not known to segregate OSD, but regarded as phenotypic dwarfs as a breed characteristic, all tested homozygous normal for COL9A3 and COL9A2 mutations.

COL9A3 Genotype / Phenotype
Normal / Retinal folds and/or vitreal membranes and normal skeleton / Dwarf
Not known to have folds / Retinal detachment, retinal folds
Homozygous mutant (-/-) / 0 / 0 / 0 / 5
Heterozygous carrier (+/-) / 7 / 1 / 0 / 0
Homozygous normal (+/+) / 81 / 21 / 2 / 1
Total / 88 / 22 / 2 / 6

COL9A2 Genotype / Phenotype
Normal / Retinal folds and/or vitreal membranes and normal skeleton / Dwarf
Homozygous mutant (-/-) / 0 / 0 / 0
Heterozygous carrier (+/-) / 2 / 0 / 0
Homozygous normal (+/+) / 59 / 0 / 0
Total / 61 / 0 / 0

Breed / Number of dogs tested for COL9A3 / Number of dogs tested for COL9A2
1 / American cocker spaniel / 2 / 5
2 / American Eskimos / 3 / 10
3 / American Pittbull terrier / 0 / 5
4 / American Staffordshire terrier / 0 / 5
5 / Australian cattle dogs / 6 / 5
6 / Basenji / 2 / 1
7 / Border Collies / 10 / 5
8 / Boxer / 1 / 1
9 / Cavalier King Charles Spaniel (a) / 2 / 2
10 / Chesapeake bay retriever / 4 / 5
11 / Chinese crested (b) / 5 / 6
12 / Collie / 5 / 0
13 / Coton de Tulear (c) / 1 / 1
14 / Doberman Pinscher (d) / 2 / 2
15 / English cocker spaniel (e) / 9 / 6
16 / English Mastiff (f) / 7 / 7
17 / English springer spaniel / 0 / 7
18 / Entelbucher mountain dogs / 1 / 7
19 / Golden Retriever (g) / 6 / 6
20 / Great Dane (h) / 1 / 0
21 / Irish Setter / 1 / 0
22 / Newfoundland (i) / 1 / 0
23 / Nova scotia duck tolling retriever (j) / 5 / 5
24 / Papillon / 0 / 7
25 / Poodle / 3 / 7
26 / Portuguese water dogs / 4 / 5
27 / Tibetan terrier / 0 / 8
Total / 81 / 118

a- both dogs have retinal folds, b- 1 of 5 has retinal folds, c- 1 dog with folds, d- 1 dog with folds, e- 1 dog with folds, f- all 7 dogs have retinal folds, g- 2 dogs with folds, h- this dog is a dwarf with retinal dysplasia, i- this dog has dwarfism but normal eye exam, j- one dog with folds.

D. Identification of 14 dogs from 5 breeds considered to be phenotypically dwarf as a breed characteristic - all tested homozygous normal for both the Col9a2 and Col9a3 OSD-associated mutations

Breed / Number of dogs
1 / American Bulldog / 2
2 / Pomeranian / 1
3 / Corgi / 4
4 / Glen of Imaal terrier / 6
5 / Dachshund / 1
Total / 14

12/9/18

Supplementary Table 3. Canine primer sequences used to amplify the complete coding region of A. COL9A3 (primer pairs 1 to 11) and B. COL9A2 (primer pairs 12 to 21) from retinal cDNA of normal and OSD-affected dogs. C. Primers used to establish the 3’ ends of the genes. D. Primers used to validate the 3’ RACE products. E. Primers used to retrieve the correct 5’ end of the COL9A2 gene

Pair / Forward primer name / Forward primer sequence / Reverse primer name / Reverse primer sequence
A.COL9A3 primers used to amplify the coding sequence.
1 / COL9A3_5UTR_F / gcgcgcagagccgctgagag / COL9A3exon8R / attccaccggggacaccact
2 / COL9A3rg1R(exon10) / ggaggacccctgggtgac
3 / COL9A3exon12R2 / cgacctctccttgatctcctt
4 / COL9A3rg1F(exon1_2) / gcagaaagtgggacctcaag / COL9A3exon8R / attccaccggggacaccact
5 / COL9A3rg1R(exon10) / ggaggacccctgggtgac
6 / COL9A3exon4F / aagccagggaagccaggaga / COL9A3exon8R / attccaccggggacaccact
7 / COL9A3rg1R(exon10) / ggaggacccctgggtgac
8 / COL9A3exon8F / agaggcaggagagagtggtgt / COL9A3exon12R / cttgccgacctctccttgat
9 / COL9A3exon25_26R / aacgccctggtctcccttag
10 / COL9A3exon20_21F / gaagggagaactgctggagag / COL9A3exon29_30R / gatttgttcgctgagcatcc
11 / COL9A3exon26F / ctggggacaaaggagagctg / COL9A3_3UTR_R2 / cccgaggtacgatgttagagc
B. COL9A2 primers used to amplify the coding sequence.
12 / COL9A2_5UTR_F4 / ccgccccgtccgagagcagc / COL9A2exon4R / gcttcccatcaggcccatctgg
13 / COL9A2exon1F / gagcctccgccgcccgcatg / COL9A2exon4_5R / gctccagttagaccatcaatcc
14 / COL9A2rg1F(exon1_2) / ctggcgcagatcagaggt / COL9A2rg1R(exon10_11) / cagttggtcggacacaagaa
15 / COL9A2exon2F / cctggatccgacggcatcgac / COL9A2exon7_8R / gtggtccaggaggtccagcaaag
16 / COL9A2exon4_5F / tgggattgatggtctaactgg / COL9A2exon22R / gggtccgatttctccttgag
17 / COL9A2exon21F / ctccctggattctctggtcc / COL9A2exon26R / gcccttctcgcctttctctc
18 / COL9A2exon24F / cttgccaggcatcaagggagac / COL9A2exon31R / ggtcacccttctctccacgtttt
19 / COL9A2exon24_25F / atcaagggagacaagggcttcc / COL9A2exon30R2 / ctcacagcgacctctgccagt
20 / COL9A2exon29F / cgaccagcacatcgtgaccgt / COL9A2exon32R / tccttgccgttgattgcctg
21 / COL9A2exon30_31F2 / ccaagggaaaacgtggagagaag / ColA2_3UTR_R2 / gaaagctggcttcctggtctgag
C. RACE-PCR primers
22 / A3_3'RACE_F1 / cggggaccgaggagacaaaggctccgc
23 / A2_3’RACE_F9 / ccgggcaggcaatcaacggcaaggacg

D. Validation of the RACE products

24 / A2_exon27_28F / ccaagggacagcaaggagtc / A2_Cds_1R / accagcagtcacccagcaagt
25 / A2_Cds_2R / accagcagtcacccagcaagtc
26 / A2_Cds_4R / tacagaaaggctggggtggatctc
27 / A3_exon26F / ctggggacaaaggagagctg / A3_3UTR_AR1 / tgttagattggggtcgctcgggtt
28 / A3_3UTR_AR2 / tatcacaacaggtaatagcagtagactttcc
29 / A2_3endgapF1 / ccccacccaccctgagat / A2_3endgapR1 / cagaacagaacaaaaggcagagg
30 / A2_3endgapF2 / ggacttgctgggtgactgct / A2_3endgapR2 / cttcctgtggggagcctgat
31 / A2_3endgapF3 / Caccttcctcccctggacttgct / A2_3endgapR3 / aaatgcttgggtctcacactgga

E. Primers used to retrieve the correct 5’ end of the COL9A2 gene

32 / COL9A2_5UTR_F2 / tgtggtaggctttcctgaaga / A2i2R2 / cccgacccaggttagagact
33 / COL9A2_5UTR_F3 / aaaagcctgacctcctagctg
34 / COL9A2_5UTR_F7 / ccggttctgtgctgctaagccag
35 / A2_exon1_F / gagcctccgccgccccgcatg / A2int1R11 / cgcctaacccgaaaggagcac
36 / A2_exon1_F2 / ctgctgctgctccaggggctc
37 / A2_F11 / gcccggttctgtgctgctaag
38 / A2_exon1_F2 / ctgctgctgctccaggggctc / A2int1R / gtgaatgggcaccattgtct
39 / A2_F10 / ccggttctgtgctgctaag / A2int1R10 / cctaacccgaaaggagcac

Supplementary Table 4. Primer sequences used to establish the Col9a2 deletion in drd2-affected dogs.

Pair / Name / Forward primer sequence / Location on Chromosome 15 (2004 assembly) / Name / Reverse primer sequence / Location on Chromosome 15 (2004 assembly) / Amplified in affected (template)
1 / ZNF643F1 / ggtgaagttgcgccatctgaa / 5542658-5542678 / ZNF643R1 / ggaggacttgctggtttctgtttt / 5533234-5533257 / Yes (cDNA)
2 / ZNF643F2 / aataggctggttggtgggaag / 5542573-5542593 / ZNF643R2 / cgggggaaaacgaaaagaaat / 5542364-5542384 / Yes (gDNA)
3 / SMAP1L_exon8F / agccagggagcaaatcagaag / 5571740-5571760 / SMAP1L_exon9R / tgggtggtcatcattccatt / 5570986-5571005 / Yes (cDNA)
4 / SMAP1L_exon3F / gatggggaatggaaaggcaaa / 5579343-5579363 / SMAP1L_exon7R / atggagcaggacacaggagca / 5572704-5572724 / Yes (cDNA)
5 / SMAP1L_intron1F / tggggagaaggggatagttcc / 5609358-5609378 / SMAP1L_intron1R / gaaacggagagttgggggtag / 5608757-5608777 / Yes (gDNA)
6 / Part1F / cccagtcgggaggtgaggac / 5610811-5610830 / Part1R / gcggaggtggagaggaagag / 5611217-5611236 / Yes (gDNA)
7 / Part2F / aagtgccaagagagggacagg / 5618340-5618360 / Part2R / aagactcaaggtggggagcag / 5618659-5618679 / Yes (gDNA)
8 / Part3F / aagaaagagaatacatcccccaat / 5622637-5622660 / Part3R / tggagaacagaacaccccagt / 5622990-5623010 / Yes (gDNA)
9 / Part4F / cgcagaacagaaggtgtgagc / 5627091-5627111 / Part4R / cagtttccacgaagccatcaa / 5627419-5627439 / Yes (gDNA)
10 / Part5F / ggaggaagaaggtgagcaagg / 5631669-5631689 / Part5R / ggcagagggaactttgtaagca / 5631994-5632015 / Yes (gDNA)
11 / Part6F / ggcagaggagaggtgataggc / 5637004-5637024 / Part6R / ttgtccagcaaggggagtgt / 5637378-5637397 / Yes (gDNA)
12 / Part7F / cagtggaaaggttgggataggag / 5637269-5637291 / Part7R / cctggttagtgttgttgctgttg / 5637773-5637795 / Yes (gDNA)
13 / Part8F / agtttgggagcacagatgatgag / 5637968-5637990 / Part8R / ccagcaagaaataggaggaggag / 5638505-5638527 / Yes (gDNA)
14 / Part9F / gtgtgtacaagtgtgcgtcttgtc / 5639826-5639849 / Part9R / cacctgggcttataacctgaacc / 5640411-5640433 / Yes (gDNA)
15 / Part10F / catctctccctcactccctcct / 5641212-5641233 / Part10R / tcacccctctcccagtctcttag / 5641693-5641715 / No (gDNA)
16 / A2int1R / gtgaatgggcaccattgtct / 5642547-5642566 / Yes, but gave a smaller product compare to Normal
17 / Part11F / gctgaccttgtggattttctcc / 5641411-5641432 / Part11R / agaccccctcatctccctctc / 5642046-5642066 / No (gDNA)
18 / A2int1R12 / gctcacggggaaccgactgtgc / 5643103-5643124 / Yes, but gave a smaller product compare to Normal
19 / A2int1R / gtgaatgggcaccattgtct / 5642547-5642566 / Yes, but gave a smaller product compare to Normal
20 / A2int1F3 / gcgagacaatggtgcccattcacg / 5642544-5642567 / A2int1R12 / gctcacggggaaccgactgtgc / 5643103-5643124 / Yes (gDNA)
21 / A2int1F4 / gcagggcacccgctcaccgtc / 5642830-5642850 / Yes (gDNA)
22 / A2intron1F2 / acagtcggttccccgtga / 5643105-5643122 / A2intron2R2 / cccgacccaggttagagact / 5643284-5643303 / Yes (gDNA)

Supplementary Table 5.Genomic primers and expected PCR product sizes, designed to identify dogs affected (homozygous mutant), carrier (heterozygous) and normal (homozygous wildtype) for OSD-associated COL9A3 (primer pairs 1-3), and COL9A2 (primer pairs 4-5) mutations. In primers 2 forward and 3 forward, the nucleotide mismatches are bold and underlined.

# / Forward primer name / Forward primer sequence / Reverse primer name / Reverse primer sequence / Expected size in wt / Expected size in carrier / Expected size in affected
1 / COL9A3test1F / gctgccactgggctcctttcttcg / COL9A3test1R / agcaggagcagggccagcgtg / 248 bp / 248 bp & 249 bp / 249 bp
2 / ASE_wt_Forward / ggcgcagccatggccgggac / ASE_reverse / ggtcagggtggcggccaggagc / 72 bp / 72 bp / No band
3 / ASE_mut_Forward / ggcgcagccatggccgggcg / ASE_reverse / ggtcagggtggcggccaggagc / No band / 73 bp / 73 bp
4 / COL9A2_Part11F / gctgaccttgtggattttctcc / COL9A2intron1R / gtgaatgggcaccattgtct / 1,445 bp / 178 bp / 178 bp
5 / COL9A2_Part10F / catctctccctcactccctcct / COL9A2_Part10R / tcacccctctcccagtctcttag / 504 bp / 504 bp / No band

Supplementary Table 6. Genotype, clinical phenotype, and multimap coding of non-dwarf Labrador colony dogs from intercrosses (A), and of non-dwarf samoyed colony dogs from intercrosses (B). nvl= no visible lesions, OD= right eye, OU= right and left eye, OS= left eye. NA= not available.

Animal / Genotype / Clinical phenotype / Multimap code for initial analysis / Multimap code for final analysis
1 / Carrier / Inconclusive / 1,2 / 1,2
2 / Normal / Inconclusive / 1,2 / 2,2
3 / Carrier / Corneal dystrophy, micropapilla, but no folds or other expected lesions / 1,2 / 1,2
4 / Carrier / Exaggerated Mittendorf dot and hyaloid remnant OS, but no folds. / 1,2 / 1,2
5 / Carrier / Vitreal strands OU / 1,2 / 1,2
6 / Carrier / Vitreal strands OS / 1,2 / 1,2
7 / Normal / Small optic nerve head OU, but no folds. / 1,2 / 2,2
8 / Carrier / No folds or other expected lesions / 1,2 / 1,2
9 / Carrier / Condensates or folds OU / 1,2 / 1,2
10 / Carrier / folds / 1,2 / 1,2
11 / Carrier / 1 fold OD / 1,2 / 1,2
12 / Carrier / multiple retinal folds / 1,2 / 1,2
13 / Normal / No overt folds - some small white spots / 1,2 / 2,2
14 / Carrier / folds / 1,2 / 1,2
15 / Normal / a few fine folds in tapetal fundus OS / 1,2 / 2,2
16 / Normal / No folds or other expected lesions / 1,2 / 2,2
17 / Carrier / multiple retinal folds / 1,2 / 1,2
18 / Carrier / Vitreal Strands, no folds / 1,2 / 1,2
19 / Carrier / No folds, but a few vitreal strands / 1,2 / 1,2
20 / Carrier / 1 fold OD, vitreal membranes OU / 1,2 / 1,2
21 / Carrier / No folds, vitreal membranes OU / 1,2 / 1,2
22 / Carrier / folds / 1,2 / 1,2
23 / Carrier / multi retinal folds OS, vitreal membranes OD / 1,2 / 1,2
24 / Carrier / Folds OS, Vitreal membranes OS / 1,2 / 1,2
25 / Carrier / multiple retinal folds and degenerate vitreous OS / 1,2 / 1,2
26 / Carrier / multiple retinal folds OU / 1,2 / 1,2
27 / Carrier / marked vitreal condensates - no folds / 1,2 / 1,2
28 / Carrier / NA / 1,2 / 1,2
29 / Carrier / NA / 1,2 / 1,2
30 / Carrier / NA / 1,2 / 1,2

Animal / Genotype / Clinical phenotype / Multimap entry for the initial analysis / Multimap entry for the final analysis
1 / Carrier / multi retinal folds / 1,2 / 1,2
2 / Carrier / one fold at first exam, nvl at second exam / not used / 1,2
3 / Normal / nvl / not used / 2,2
4 / Carrier / multi retinal folds / 1,2 / 1,2
5 / Carrier / 1 fold OD, vitreous degenerate OD / 1,2 / 1,2
6 / Carrier / multi retinal folds OU, vitreous degenerate OU / 1,2 / 1,2
7 / Carrier / folds / 1,2 / 1,2
8 / Carrier / 1 fold OU / 1,2 / 1,2
9 / Carrier / folds / 1,2 / 1,2
10 / Carrier / multi retinal folds / 1,2 / 1,2
11 / Carrier / multi retinal folds / 1,2 / 1,2
12 / Carrier / inconclusive / 1,2 / 1,2
13 / Carrier / nvl / 1,2 / 1,2
14 / Carrier / multi retinal folds / 1,2 / 1,2
15 / Carrier / multi retinal folds OS / 1,2 / 1,2
16 / Carrier / retinal folds and degenerate vitreous / 1,2 / 1,2
17 / Carrier / No folds but degenerate vitreous / 1,2 / 1,2
18 / Carrier / multiple retinal folds and degenerate vitreous / 1,2 / 1,2
19 / Carrier / multi retinal folds / 1,2 / 1,2
20 / Carrier / multi retinal folds / 1,2 / 1,2
21 / Carrier / multi retinal folds / 1,2 / 1,2
22 / Carrier / folds / 1,2 / 1,2
23 / Carrier / folds at first exam, nvl at second exam / 1,2 / 1,2
24 / Normal / No folds seen / 2,2 / 2,2
25 / Carrier / folds OS / 1,2 / 1,2
26 / Carrier / No folds but degenerate vitreous / 1,2 / 1,2
27 / Normal / No folds seen / 2,2 / 2,2
28 / Carrier / No folds seen / 2,2 / 1,2
29 / Normal / No folds seen / not used / 2,2

12/9/18

Supplementary figure 1. Canine COL9A3 and COL9A2 sequences of normal and mutated alleles.

Sequence of the COL9A3 mRNA in normal dogs. Upper case indicates coding sequence, boxed are the first methionine and the stop codon, SNPs are in bold, underlined and italic are the polyadenylation signals, * indicates the alternative poly A tail location
Sequence of the canine COL9A3 mRNA in drd1-affected dogs. Upper case indicates coding sequence, boxed is the first methionine followed by uppercased-italic-five-G stretch representing the mutation. The putative stop codon in the mutated COL9A3 is bold and underline (TGA). For comparison, the normal stop codon is boxed (TAA). SNPs are in bold.
Sequence of the canine COL9A2 mRNA gene. Upper case indicates coding sequence, boxed are the first methionine and the stop codon, SNPs are in bold, underlined and italic is the polyadenylation signal.
Sequence of the 5’ end of canine COL9A2 gene. Italic and bold indicates the sequence deleted in drd2-affected dogs, Upper case indicates coding sequences of exon 1 and 2, boxed is the first methionine.

cgcgggcgcagccATGGCCGGGGCCCCCACGCTGGCCCTGCTCCTGCTCGCGCGGCTCCTGGCCGCCACCCTGACCGGGACCGGGGCGCAGAAAGTGGGACCTCAAGGCCCCCCCGGTCCCCAAGGGCCACCTGGGAAGCCGGGCAAGGATGGCATTGATGGAGAAGCTGGGCCTCCCGGTCTGCCCGGGTCCCCGGGACCAACAGGGGCCCCAGGGAAGCCAGGGAAGCCAGGAGAGGCCGGGCTGCCRGGACTGCCCGGTGTGGACGGCCTGACGGGGCAGGACGGACCTCCCGGACCCAAGGGCGCACCTGGGGAACRGGGAAGCCTGGGACCCCCGGGGCCTCCCGGGTTGGGGGGCAAAGGCCTCCCCGGACCCCCCGGAGAGGCAGGAGAGAGTGGTGTCCCCGGTGGAATCGGCCTCCGGGGCCCCCCGGGACCCTCTGGACTGCCAGGCCTCCCCGGCCCCCCCGGACCTCCCGGACCCCCCGGTCACCCAGGGGTCCTCCCTGAGGGCGCCACTGACCTTCAGTGCCCGGCCATCTGCCCACCAGGCCCCCCCGGTCCCCCAGGAATGCCGGGGTTCAAGGGGCCCACCGGCTACAAAGGAGATCAAGGAGAGGTCGGCAAGGACGGCGAGAAGGGCGATCCTGGCCCCCCTGGGCCCGCTGGCGTCCCTGGCTCTGTGGGGCTGCAGGGGCCTCGGGGACTCCGAGGTCTGCCCGGGCCGGCTGGGCCCCCAGGGGATCGGGGTCCCATCGGATTCCGAGGGCCGCCAGGGATCCCAGGAGCCCCCGGGAAAGTGGGTGACAGAGGCGAGAGGGGCCCAGAGGGTTTCCGCGGCCCCAAGGGTGACCTTGGCAGACCCGGCCTCAAGGGAGTCCCCGGGATGGCCGGGCCGGGCGGGGAGCCGGGAATGCCAGGCAAGGACGGCCGGGATGGCGTGCCGGGACTGGACGGCGAGAAGGGAGAGGCCGGTCGCAACGGTGCCCCAGGAGAGAAGGGTCCCAACGGGCTGCCGGGCCTCCCGGGTCGAGCAGGGTCCAAGGGCGAGAAGGGAGAACTGGGCCGAGCTGGAGAGCTGGGGGAGGCTGGCCCCTCGGGAGAGCCTGGCATCCCGGGGGACGTTGGCGTGCCTGGGGAGCGTGGTGAGGCTGGCCACAGGGGCTCGGCGGGGGCTCTGGGCCCACAAGGCCCTCCTGGAGCCCCTGGCGTCCGCGGCTTCCAGGGCCAGAAGGGCAGCATGGGCGACCCCGGCCTGCCGGGTCCCCAGGGCCTCCGAGGTGCCTCAGGTGACCGGGGCCCGGGGGGAGCCGCAGGCCCTAAGGGAGACCAGGGCGTTGCAGGTTCCGACGGCCTCCCTGGGGACAAAGGAGAGCTGGGTCCCGGTGGCCCGGTCGGACCCAAAGGAGAGGCTGGCAGTCGAGGGGAGCTGGGCCCCAAGGGCATCCAGGGTCCCAAYGGCACCAGCGGCGTCGAGGGCCTCCCGGGCCCGCCCGGCCCCGTGGGCTTCCCGGGCGTCCAGGGCGTGCCCGGCATCACCGGGAAACCGGGGGTTCCGGGGCGAGAAGCCAGCGAGCAGCACATCCGGGAGCTGTGCGGGGGGATGCTCAGCGAACAAATCGCGCAGTTGGCCGCTCACCTGAGGAAGCCTCTGGCGCCCGGATCCGCCGGGCGGCCTGGGCCAGCGGGGCCCCCAGGCCCCCCGGGGCCCCCAGGCTCCATCGGCCACCCCGGTGCCCGAGGGCCCCCTGGATACCGCGGCCCCACCGGAGAGCTGGGGGACCCGGGGCCCAGAGGGGCCCCCGGGGACCGAGGAGACAAAGGCTCCGCRGGCGCGGGTCTGGACGGGCCGGCCGGGGACCAGGGCCTCCAAGGACCACAAGGCGTGCCTGGCGTTAGCAAAGACGGCCGCGACGGGGCCAACGGCGAGCCCGGGCCTCCAGGCGATCCTGGCCTCCCCGGTGCTGTGGGTGCTCAGGGGACACCCGGCATCTGCGACACCTCGGCCTGCCAAGGAGCTGTGATGGGAGGCGGCGGGGAAAAGTCAGGTTCTAGAAGCTTCTAAaacgtaaccaaaggaactgaggtggtggtgccgatgccagcagggagctctgragggacgccaggagtcggcggcrctctgtgccrggaggggacagcatgccacccgcggcrgcccccgcggccctcctggcctgcccctcgttcctcggtggcagcagccggacacttgtccctgagagcatccgtcatagacctgttgcccagtgtygtcctccgygcatctccacctgtgcatcagctctaacatcgtacctcgggcacgggtcccgtgtgcacgaccgagaccccacttctgagagggagacagggcacaggtacacaaagtcacgtacgattccacgagacctaaaaccgcaagctgttcacacagcgcttgtgtaaggagrtctgctcacaggtagcggcagatgaacccgagcgaccccaatctaacaaataaaggatttcacgtacggt*accccctccacacatggtctagtctgatttacctgcttcctgggtaaaaacttttgactgtctctgagtaacagtatttattgaaggaaagtctactgctattacctgttgtgataaataaagccactcgtgtaaaaaaaaaaaaaaaaaaaaaaa

cgcgggcgcagccATGGCCGGGGGCCCCCACGCTGGCCCTGCTCCTGCTCGCGCGGCTCCTGGCCGCCACCCTGACCGGGACCGGGGCGCAGAAAGTGGGACCTCAAGGCCCCCCCGGTCCCCAAGGGCCACCTGGGAAGCCGGGCAAGGATGGCATTGATGGAGAAGCTGGGCCTCCCGGTCTGCCCGGGTCCCCGGGACCAACAGGGGCCCCAGGGAAGCCAGGGAAGCCAGGAGAGGCCGGGCTGCCGGGACTGCCCGGTGTGGACGGCCTGACGGGGCAGGACGGACCTCCCGGACCCAAGGGCGCACCTGGGGAACGGGGAAGCCTGGGACCCCCGGGGCCTCCCGGGTTGGGGGGCAAAGGCCTCCCCGGACCCCCCGGAGAGGCAGGAGAGAGTGGTGTCCCCGGTGGAATCGGCCTCCGGGGCCCCCCGGGACCCTCTGGACTGCCAGGCCTCCCCGGCCCCCCCGGACCTCCCGGACCCCCCGGTCACCCAGGGGTCCTCCCTGAGGGCGCCACTGACCTTCAGTGCCCGGCCATCTGCCCACCAGGCCCCCCCGGTCCCCCAGGAATGCCGGGGTTCAAGGGGCCCACCGGCTACAAAGGAGATCAAGGAGAGGTCGGCAAGGACGGCGAGAAGGGCGATCCTGGCCCCCCTGGGCCCGCTGGCGTCCCTGGCTCTGTGGGGCTGCAGGGGCCTCGGGGACTCCGAGGTCTGCCCGGGCCGGCTGGGCCCCCAGGGGATCGGGGTCCCATCGGATTCCGAGGGCCGCCAGGGATCCCAGGAGCCCCCGGGAAAGTGGGTGACAGAGGCGAGAGGGGCCCAGAGGGTTTCCGCGGCCCCAAGGGTGACCTTGGCAGACCCGGCCTCAAGGGAGTCCCCGGGATGGCCGGGCCGGGCGGGGAGCCGGGAATGCCAGGCAAGGACGGCCGGGATGGCGTGCCGGGACTGGACGGCGAGAAGGGAGAGGCCGGTCGCAACGGTGCCCCAGGAGAGAAGGGTCCCAACGGGCTGCCGGGCCTCCCGGGTCGAGCAGGGTCCAAGGGCGAGAAGGGAGAACTGGGCCGAGCTGGAGAGCTGGGGGAGGCTGGCCCCTCGGGAGAGCCTGGCATCCCGGGGGACGTTGGCGTGCCTGGGGAGCGTGGTGAGGCTGGCCACAGGGGCTCGGCGGGGGCTCTGGGCCCACAAGGCCCTCCTGGAGCCCCTGGCGTCCGCGGCTTCCAGGGCCAGAAGGGCAGCATGGGCGACCCCGGCCTGCCGGGTCCCCAGGGCCTCCGAGGTGCCTCAGGTGACCGGGGCCCGGGGGGAGCCGCAGGCCCTAAGGGAGACCAGGGCGTTGCAGGTTCCGACGGCCTCCCTGGGGACAAAGGAGAGCTGGGTCCCGGTGGCCCGGTCGGACCCAAAGGAGAGGCTGGCAGTCGAGGGGAGCTGGGCCCCAAGGGCATCCAGGGTCCCAATGGCACCAGCGGCGTCGAGGGCCTCCCGGGCCCGCCCGGCCCCGTGGGCTTCCCGGGCGTCCAGGGCGTGCCCGGCATCACCGGGAAACCGGGGGTTCCGGGGCGAGAAGCCAGCGAGCAGCACATCCGGGAGCTGTGCGGGGGGATGCTCAGCGAACAAATCGCGCAGTTGGCCGCTCACCTGAGGAAGCCTCTGGCGCCCGGATCCGCCGGGCGGCCTGGGCCAGCGGGGCCCCCAGGCCCCCCGGGGCCCCCAGGCTCCATCGGCCACCCCGGTGCCCGAGGGCCCCCTGGATACCGCGGCCCCACCGGAGAGCTGGGGGACCCGGGGCCCAGAGGGGCCCCCGGGGACCGAGGAGACAAAGGCTCCGCAGGCGCGGGTCTGGACGGGCCGGCCGGGGACCAGGGCCTCCAAGGACCACAAGGCGTGCCTGGCGTTAGCAAAGACGGCCGCGACGGGGCCAACGGCGAGCCCGGGCCTCCAGGCGATCCTGGCCTCCCCGGTGCTGTGGGTGCTCAGGGGACACCCGGCATCTGCGACACCTCGGCCTGCCAAGGAGCTGTGATGGGAGGCGGCGGGGAAAAGTCAGGTTCTAGAAGCTTCTAAaacgtaaccaaaggaactgaggtggtggtgccgatgccagcagggagctctggagggacgccaggagtcggcggcactctgtgccgggaggggacagcatgccacccgcggcagcccccgcggccctcctggcctgcccctcgttcctcggtggcagcagccggacacttgtccctgagagcatccgtcatagacctgttgcccagtgtcgtcctcc

ccgcgccgcccgcccgcccgagcctccgccgcccgcATGGCCGCCGCCCGCCGCCTCCTGCTGCTGCTCCAGGGGCTCGCGCTCGCCCTGGCGCAGATCAGAGGTCCGCCCGGAGAACCGGGGCCCCCGGGCCCCCCGGGGCCGCCGGGAGTGCCTGGATCCGACGGCATCGACGGTGACAAGGGGYCCCCTGGGAAAGCCGGCCCTCCGGGACTGAAGGGAGAGCCTGGCAAACCCGGGCCAGATGGGCCTGATGGGAAGCCTGGGATTGATGGTCTAACTGGAGCCAAGGGGGAGCCTGGCCCCATGGGGATCCCTGGAGTCAAGGGCCAGCCTGGGCTCCCCGGTCCCCCTGGCCTGCCGGGCCCTGGCTTTGCTGGACCTCCTGGACCACCTGGACCTGTTGGCCTCCCCGGTGAGATTGGAATCACAGGCCCCAAGGGGGATCCCGGACCAGATGGGCCATCGGGGCCCCCGGGGCCACCTGGCAAACCGGGCCGACCCGGAACCATCCAGGGCCTGGAAGGCAGCGCGGATTTCTTGTGTCCGACCAACTGTCCGGCAGGGGTGAAAGGCCCCCCAGGGCTGCAGGGGGTGAAGGGGCATCCTGGCAAACGCGGGGTTCTGGGCGATTCTGGCCGCCAGGGGAAGCCGGGTCCCAAGGGAGATGTGGGTGCCTCTGGAGAGCAAGGCATCCCTGGACCACCGGGTCCCCAGGGCATCAGGGGCTACCCGGGCATGGAGGGACCCAAAGGAGAGATGGGTCCTCGTGGGTACAAAGGCATGGTGGGCTCCATTGGTGCTGCCGGGTCACCCGGTGAGGAAGGTCCACGGGGGCCACCGGGCCGAGCGGGGGAGAAGGGCGATGTGGGTGGCCAAGGTCTCCGAGGACCTCAGGGCATAACAGGCCCGAAGGGAGCAACCGGGCCCCCAGGCATCGATGGCAAGGATGGGACCCCAGGCACGCCAGGCATGAAGGGCAGTGCGGGACAGGCAGGGCGGCCAGGAAACCAAGGCCACCAAGGCCTAGCGGGCGTGCCGGGCCAGCCTGGGACAAAAGGAGGCCCGGGAGACAAGGGTGAACCAGGCCAGCAGGGCCTCCCTGGATTCTCTGGTCCCCCTGGGAAGGAGGGAGAGCCAGGACCTCAAGGAGAAATCGGACCCCGAGGCATCATGGGGCAGAAGGGTGACCAGGGTGAGAGGGGGCCGGTGGGGCAGCCAGGCCCTCAGGGACGGCAGGGCCCCAAGGGGGAGCAGGGCCCCCCCGGAATTCCAGGGCCCCAAGGCTTGCCAGGCATCAAGGGAGACAAGGGCTTCCCGGGGAAGACCGGCCCCCGCGGCAGCGTGGGCGACCCGGGGGTGGCCGGCCTCCGGGGAGAGAAAGGCGAGAAGGGCGAGTCCGGCGAGCCGGGGCCCAAGGGACAGCAAGGAGTCCGCGGAGAAGCCGGCTACCCGGGCCCCAGCGGGGATGCGGGCGCCCCGGGGGTGCAGGGCTACCCCGGGCCCCCCGGCCCTCGAGGACTGGCGGGAGACCGAGGCGTGCCCGGACTGCCCGGGAGACAGGGCGTGGCGGGCCGGGACGCCAGCGACCAGCACATCGTGACCGTGATGATGAAGATGATGCAAGAGCAACTGGCAGAGGTCGCTGTGAGTGCCAAGCGGGAGGCCCTGGGTGCCATCGGGATGGTGGGTCCGCCAGGACCCCCCGGGCCTCCTGGGTACCCAGGCAAGCAGGGACCTCATGGGCACCCTGGCCCTCGGGGCGTTCCTGGCATCGTGGGAGCCGTGGGTCAGATTGGCAACACCGGCCCCAAGGGAAAACGTGGAGAGAAGGGTGACCGGGGAGAGATGGGACGCGGGCACCCCGGGATGCCTGGGCCCCCCGGGATCCCAGGGCTCCCCGGCCGGCCCGGGCAGGCAATCAACGGCAAGGACGGGGCCCGCGGCGCCCCAGGGGCCCCGGGGGAAGCAGGCCGACCGGGTCCGCCGGGCCCCGCGGGGCTGCCCGGCTTCTGTGAGCCCGCGGCCTGCCTGGCAGCCTCGGCCTACGCCTCTGCGCGCCTCACAGAGCCCGGATCCATCAAAGGGCCATGAgcaggaggccaggacagagcctggcgggcatcctggggggtggggggggactagattccagcggggtggacacgcaccccatccctcagaccaggaagccagctttccagacccttccatccgggaccctggagtcctgaggaaaaggaaattctaagattcgggggaagcgaagggagggcattgggatgaaaaggtgaggccaacagacggcaagggtcactggagagtcccagctccaaagaggtggctttcctcccagagcacgactcacaaggtaccaaagcaaatgtctccgtaatctgtgctgtcctccagtggccacctctttggggcaggtggactgaggtctctgccggtccctgagcccaccttcctcccctggacttgctgggtgactgctggtgggtgacccaccccgaggtggccgtctgaagcggccccactacttcctccctgcgtccctcccatctgagtatttaagcacccctcccccttctcttcctggccttacccccacccaccctgagatccaccccagcctttctgtaaataaactccccccccgccccgacttgggtacaaacaaaaaaaaaaaaaaaaaa

1 cctgtcagct tacttcctat aaactccaaa ttcctcctcg gtccattagg cagagcaatc

61 aaattctccc aaattcagct ccaggaacac ctctcaaagc ccagggccct ttgctcacaa

121 tgccacgcgg ggttggactt tcccaggctc cagaggggtt tctcctggac tattttagct

181 gaccttgtgg attttctccc ccatgctgggcacaggactg gtaaacaaag tatattctcc

241 ataaatcata atgaaagctg aaatttatga agcccctact acttcatatg aagtacttag

301 catcatctca ttcaatcttc acaaaaattc tataagatag gattattatt atcccctttt

361 acagaggagg acactatatg gtttcaggtg cctgtggtca ccgttagcga cagattccaa

421 agcacttcac caacattgct taagggaagg aaaagggagc taagagactg ggagaggggt

481 gatgctggac aaacggggga gggggagtgc ttcatggagg aggtcacccc gagggatacc

541 tagagactct ggtgcttcaa gaagctgcag tgaagaaaag gcaaggtgag gagaaggggg

601 aggaggagga ggaggaagag gaggaggtgg aggaggagga ggaggacgct ggacggtggg

661 ggcctgggca gggaggcttc aaggagagca tggccaaggg cggctccagg agcaacagga

721 gcttcagtgc ttccgggggc actaagaggg cctggggctc gctggtggcc tccaaaagcc

781 tgacctccta gctgcggcgg gagcagggag gtgagaggga gatgaggggg tctggaggtg

841 cggcacgggg tgcatggggg ggctcccggg accgccgacc cgtaaggctt gtggtaggct

901 ttcctgcaga agaggcctgc cggtctgtgc tgctaagcca ggtgggtggg ggcctctgcg

961 aactggggtg caggacccgg gcggtggggg cctctgcgac ctggggtgca ggacccacgg

1021 ctcgctctcg gcctgcgccc tggagcagag gagcgacggg ccgaggctcg gcccggggtc

1081 aggctgcagg ggcccccccg acggccggcg cctcccctcg ggggcagagg gtgggcgctg

1141 gctgcggcgc cgggcggggc ggagctgcgc gggcctcggc ccctcggccg ccctctcggg
1201 tgacggcccc gcggcccccg ccccgccccg ccgcatattc aggcgcccgc ccgcccgccc

1261 cgtccgagag cagcccgcgc cgcccgcccg cccgagcctc cgccgcccgc ATGGCCGCCG
1321 CCCGCCGCCT CCTGCTGCTG CTCCAGGGGC TCGCGCTCGC CCTGGCGCAG ATCgtaagtg

1381 ccgcggccgc cgcgcggggt cggggtgcgg ggggcggggg cggggacggg tcggggtgcg

1441 ggcctcggcg cttcccctcg gcggggcgcacagcgggcgc gtccgggcgc cgcgagggag

1501 gcggaggggc ccggggagcc aggggggccc tgtccggagg cgcgcggccc ggcctgggac

1561 aggcggcgcg gccgggagcc ccgagagccc gggtggcggg cgagacaatg gtgcccattc

1621 acggggctcc ggtgcccsgg cgccgcctcc ccccgggggg ctccagccag acgccgctcg

1681 gccccgctgc cgggggccac cggggcgcgg ggacccaccg ggaccggccc gggcgccaag

1741 cctggcacct cgctgcgcgg agggcgcggg cccggggcgg gggggggggc tgccccgcgg

1801 agggcaagcc aacgggaaac ggaggggctc cggggagggg cgccccccgc gtgctccttt

1861 cgggttaggc gcgccaggcg cagggggcag ggcacccgct caccgtcggt acctgtcgtg

1921 cgagcgagcg gtcccggagc ccgccgtcgc ctgcccgctg ggacccgcgg cgccccrgtc

1981 tcccgctccg tccaccgccc tccgccctcc gccctccgcc ctccgcgacc gcgcacgggc

2041 cccgcgggcc agcgcaccgg gacgcgctcc gcgctcgctc tccgggagcc gggtcccccc

2101 gcggccaacc cggggccgcg cctcgggtcc tgagcgccgg gaggggcacc cagggcgccg

2161 cacagtcggt tccccgtgag cgcgccgcgc cccggggggg ctctccgcag ggacccccgc

2221 cccctgcacc gtctgagggc ctccccctgt gtgtgttctc agAGAGGTCC GCCCGGAGAA

2281 CCGGGGCCCC CGGGCCCCCC GGGGCCGCCG GGAGTGCCTG GATCCGACGG CATCGACgt

Supplementary figure 2: COL9A3 and COL9A2 screening test.

COL9A3 screening test: A.1. and A.2. are the normal and affected sequences, respectively, amplified by the underlined primers. Bolded are the SNPs, boxed is the first methionine, bolded, underlined and italic are the five-G stretch presenting the mutation in drd1-affected dogs. These PCR products were sent to sequence and chromatograms were evaluated for the present or absent of the mutation. A.3 and A4 are the normal and affected sequences, respectively, amplified by the underlined primers in the allele-specific extension assay. In bold are the primers that were altered from the genomic sequence. The PCR products were ran on a 6% PAGE and evaluated for present or absent of the amplicon as an indication of the present or absent of the mutation.
B. COL9A2 test: Single underlined primers are primers that will amplify only normal allele (504 bp), double underlined primers will amplify 178 bp affected allele and 1,445 bp normal allele, italic and bold is the sequence deleted in drd2-affected chromosomes, upper case letter sequence is the coding sequences, boxed is the first metionine.

A.1.

gctgccactgggctcctttcttcgccggcgcggcgcggggcgggggcgggcggaagccccaggtgggcccggctgaatggggggcttgtgcgcgcrgggcgggacctggccgggggcccgcgccrcccgccgccccgccygtccgcccgagccccggcgccccagccccgccgcccagaggcgcgcagagccgctgagagcgcggGCGCAGCCATGGCCGGGGCCCCCACGCTGGCCCTGCTCCTGCT

A.2.

gctgccactgggctcctttcttcgccggcgcggcgcggggcgggggcgggcggaagccccaggtgggcccggctgaatggggggcttgtgcgcgcagggcgggacctggccgggggcccgcgccgcccgccgccccgcccgtccgcccgagccccggcgccccagccccgccgcccagaggcgcgcagagccgctgagagcgcggGCGCAGCCATGGCCGGGGGCCCCCACGCTGGCCCTGCTCCTGCT

A.3.

gGCGCAGCCATGGCCGGGaCCCCCACGCTGGCCCTGCTCCTGCTCGCGCGGCTCCTGGCCGCCACCctgacc

A.4.

gGCGCAGCCATGGCCGGGcGCCCCCACGCTGGCCCTGCTCCTGCTCGCGCGGCTCCTGGCCGCCACCctgacc

catctctccctcactccctcctcctgtcagcttacttcctataaactccaaattcctcct

cggtccattaggcagagcaatcaaattctcccaaattcagctccaggaacacctctcaaa

gcccagggccctttgctcacaatgccacgcggggttggactttcccaggctccagagggg

tttctcctggactattttagctgaccttgtggattttctcccccatgctgggcacaggac

tggtaaacaaagtatattctccataaatcataatgaaagctgaaatttatgaagccccta

ctacttcatatgaagtacttagcatcatctcattcaatcttcacaaaaattctataagat

aggattattattatccccttttacagaggaggacactatatggtttcaggtgcctgtggt

caccgttagcgacagattccaaagcacttcaccaacattgcttaagggaaggaaaaggga

gctaagagactgggagaggggtgatgctggacaaacgggggagggggagtgcttcatgga

ggaggtcaccccgagggatacctagagactctggtgcttcaagaagctgcagtgaagaaa

aggcaaggtgaggagaagggggaggaggaggaggaggaagaggaggaggtggaggaggag

gaggaggacgctggacggtgggggcctgggcagggaggcttcaaggagagcatggccaag

ggcggctccaggagcaacaggagcttcagtgcttccgggggcactaagagggcctggggc

tcgctggtggcctccaaaagcctgacctcctagctgcggcgggagcagggaggtgagagg

gagatgagggggtctggaggtgcggcacggggtgcatgggggggctcccgggaccgccga

cccgtaaggcttgtggtaggctttcctgcagaagaggcctgccggtctgtgctgctaagc

caggtgggtgggggcctctgcgaactggggtgcaggacccgggcggtgggggcctctgcg

acctggggtgcaggacccacggctcgctctcggcctgcgccctggagcagaggagcgacg

ggccgaggctcggcccggggtcaggctgcaggggcccccccgacggccggcgcctcccct

cgggggcagagggtgggcgctggctgcggcgccgggcggggcggagctgcgcgggcctcg

gcccctcggccgccctctcgggtgacggccccgcggcccccgccccgccccgccgcatat
tcaggcgcccgcccgcccgccccgtccgagagcagcccgcgccgcccgcccgcccgagcc

tccgccgcccgcATGGCCGCCGCCCGCCGCCTCCTGCTGCTGCTCCAGGGGCTCGCGCTC
GCCCTGGCGCAGATCgtaagtgccgcggccgccgcgcggggtcggggtgcggggggcggg

ggcggggacgggtcggggtgcgggcctcggcgcttcccctcggcggggcgcacagcgggc

gcgtccgggcgccgcgagggaggcggaggggcccggggagccaggggggccctgtccgga

ggcgcgcggcccggcctgggacaggcggcgcggccgggagccccgagagcccgggtggcg

ggcgagacaatggtgcccattcac