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. 2019 Oct 21;51(1):141–146. doi: 10.1111/age.12865

Identification of whole‐genome significant single nucleotide polymorphisms in candidate genes associated with body conformation traits in Chinese Holstein cattle

Zhengui Yan 1, Zhonghua Wang 1, Qin Zhang 1, Shujian Yue 1, Bin Yin 1, Yunliang Jiang 1, Kerong Shi 1,✉
PMCID: PMC7003999  PMID: 31633203

GWAS has been proven to be a powerful tool for detecting genetic variants associated with economically important traits such as production,1, 2 body conformation3, 4 and reproduction traits,5 and diseases.6 The objective of this study was to identify SNPs with significant association effects on body conformation traits in Chinese Holstein through the GWAS approach.

The experimental population consisted of 445 Chinese Holstein cows, with parity no. 2, raised at four farms in Shandong province of China. Hair follicle samples were collected individually within four days in the same season. In total, 29 body conformation traits were analyzed by GWAS. There were 23 phenotypes measured individually on 1–9 scores, which were body height, height at front end, body depth, chest width, stature, angularity, rump angle (RA), pin width, loin strength, foot angle, heel depth, bone quality, rear legs – side view, rear legs – rear view, udder depth, udder texture, median suspensory, fore udder attachment, front teat placement, attachment height, attachment width (AW), rear teat placement and teat length. The remaining six functional body conformation traits, measured on 1–100 scores, were stature score, rump system score, feet and legs score, udder system score, mammary system score and conformation final score (FS). The definitions of these conformation traits for cattle are available on the website of Canadian Dairy Network (https://www.cdn.ca/articles.php). The summary statistics of these phenotypes are listed in Table S1.

Each experimental animal was genotyped by the GGP BovineLD V3 SNP chip, containing 26 151 public SNPs. Because all of the animals in these study populations were female, the SNPs from the X chromosome were also counted. The data quality control procedure excluded individuals with more than 10% missing genotypes and SNPs with a call rate of less than 90%, MAF less than 3% or P‐value of the Hardy–Weinberg Equilibrium test less than 1.0E − 6. Finally, 421 animals with 20 632 SNP genotypes were retained for the subsequent GWAS analysis. Physical map length, the number of SNPs and SNP density on each chromosome based on reference genome UMD3.1, before and after the data cleaning procedure, are shown in Table S2. Results of pair‐wise LD analysis of the 421 animals from four different farms showed high similarity of LD patterns among the whole population, indicating that similar breeding histories were shared by the four subpopulations (data not provided).

A fixed‐effects linear regression model was used to carry out the GWAS according to a previous method.2, 7 Bonferroni correction for the genome‐wide significance and suggestive thresholds were computed to be 2.42E − 06 (= 0.05/20 632) and 4.85E − 05 (= 1/20 632), respectively.

The GWAS based on the mixed‐effect linear regression model identified 79 SNPs with genome‐wide significant (P < 0.05) association effects on 14 body conformation traits (Table 1, Table S3). Of these SNPs, 28 SNPs were found to be associated with RA, and 14 SNPs associated with FS (Table 1, Fig. S1). We also found six single SNPs that were associated with multiple body conformation traits of dairy cow. These SNPs were BovineHD0600000461, BovineHD1900007686, ARS‐BFGL‐NGS‐41612, BovineHD1700010514, BovineHD3000037672 and ARS‐BFGL‐NGS‐109467 (their detailed information is provided in Table S4). In this study, the DNA region from 44.6 to 65.4 Mb of chromosome 18 was identified to be associated with four different body conformation traits (AW, FS, mammary system score and RA) of dairy cows (Table 2, Table S3). This coincided with previously reported QTL or DNA regions associated with calving performance and udder traits of dairy cattle.8, 9, 10 Among these SNPs, Hapmap57004‐rs29011610, located near MON1B (MON1 secretory trafficking family member B) gene, were also found to be associated with length of productive life in Holstein cows.11 ARS‐BFGL‐NGS‐116541, which was located within the LIG1 (Ligase I) gene region, was reported to be associated with body weight at birth.12 Similarly, DNA regions on chromosomes 21, 25 and X were also identified to be significantly associated with multiple body conformation traits (Table 2, Table S3). In particular, six SNPs in the small region from 57.55 to 57.62 Mb on the chromosome 21 were found to be associated with AW and FS in the study, and these SNPs were all located within or near the gene region of SLC24A4 (Table S5). Interestingly, SLC24A5 and SLC19A2, members of solute carrier family proteins with SLC24A4, were also found to be associated with RA and FS, respectively (Table 1). Additionally, UA‐IFASA‐6670, located within the region of GABARAPL1 [GABA (A) receptor‐associated protein like 1] gene, were found to be significantly associated with median suspensory in this study. It was reported previously to affect udder attachment and height.13, 14 Also, BovineHD2200013812 was identified to have genetic effects on RA. This SNP was located within the CACNA1D (calcium channel alpha 1D subunit) gene region. Coincidentally, this SNP was also reported to be associated with the length of productive life and udder cleft.14

Table 1.

The significant SNPs and genes that are associated genome‐wide with body conformation traits

Category Trait Chromosome 1 Position SNP name P‐Value Nearest gene Distance 2 Reference Alteration MAF
Dairy strength Angularity 30 110222328 BTA‐116883‐no‐rs 1.56E − 06 LOC786124 D:27320 G A 0.2404
30 116273044 BovineHD3000032546 6.55E − 07 LOC537655 Within C A 0.3061
30 132705219 BovineHD3000037672 5.10E − 08 LOC786725 U:54284 T C 0.449
Stature 10 90906002 Hapmap39512‐BTA‐79353 5.11E − 07     T G 0.2744
19 26049717 BovineHD1900007686 2.15E − 06 AIPL1 D:7711 T C 0.03628
Chest width 17 38504014 BovineHD1700010514 1.34E − 07 LOC512119 D:176780 T C 0.09545
Body height 11 80730546 ARS‐BFGL‐NGS‐41612 4.93E − 07 KCNS3 U:112015 G A 0.1485
11 105064528 BovineHD1100030541 1.50E − 06 LOC789076 U:83296 C T 0.4728
13 21301875 BovineHD1300006183 4.26E − 07 LOC524240 Within G A 0.3515
23 39248351 BovineHD2300011340 2.39E − 06 NHLRC1 D:7508 T C 0.4341
27 39335460 Hapmap38550‐BTA‐98603 1.66E − 06 LRRC3B D:101345 C A 0.3497
Stature score 10 45053776 BovineHD1000013564 9.37E − 07 NID2 D:67117 G T 0.2324
17 38504014 BovineHD1700010514 2.42E − 08 LOC512119 D:176780 T C 0.09545
Mammary system Attachment width 9 20356212 BTA‐85319‐no‐rs 7.16E − 07 BCKDHB D:21051 T C 0.2614
18 65402237 BovineHD1800019049 4.75E − 07 LOC789374 Within C T 0.2143
19 24263948 BovineHD1900006998 1.35E − 06 LOC786649 U:52976 C T 0.4932
21 57583470 BovineHD2100016545 2.26E − 06 SLC24A4 U:12991 A G 0.4478
21 57584406 BovineHD2100016546 2.33E − 06 SLC24A4 U:12055 A C 0.4477
21 57587712 BovineHD2100016549 2.26E − 06 SLC24A4 U:8749 C T 0.4478
21 57620878 BovineHD2100016561 9.13E − 07 SLC24A4 Within G A 0.2823
21 57623572 BovineHD2100016563 9.13E − 07 SLC24A4 Within G A 0.2823
26 37203584 ARS‐BFGL‐NGS‐8275 8.12E − 07 LOC531271 Within A G 0.2381
Median suspensory 5 100206147 UA‐IFASA‐6670 6.37E − 07 GABARAPL1 Within C T 0.398
9 93370986 BovineHD0900026424 5.03E − 07 NOX3 U:2759 T C 0.325
17 73901259 BovineHD1700021616 9.77E − 07 LOC531152 D:1528 T C 0.1462
30 138304543 BovineHD3000039710 5.31E − 07 LOC782196 U:18403 T C 0.213
Mammary system score 13 28331553 ARS‐BFGL‐NGS‐109467 5.49E − 07 SEPHS1 D:16954 A G 0.4388
15 43538866 ARS‐BFGL‐NGS‐115625 2.03E − 06 SWAP70 Within G A 0.4467
18 55956772 ARS‐BFGL‐NGS‐60829 6.86E − 07 NUCB1 D:1160 C T 0.4172
30 132705219 BovineHD3000037672 7.03E − 08 LOC786725 U:54284 T C 0.449
Rear teat placement 9 25459892 BTA‐83107‐no‐rs 1.10E − 06 MIR2284O U:14492 A G 0.3307
Udder depth 25 35623801 ARS‐BFGL‐NGS‐28167 2.16E − 06 CUX1 D:12155 A G 0.4023
25 36266951 BovineHD2500010029 1.27E − 07 LOC100298352 Within T C 0.356
Udder system score 6 1770665 BovineHD0600000461 1.03E − 07 1‐Mar Within G A 0.39
Rump structure Loin strength 5 61620118 BovineHD0500017277 2.90E − 07 NEDD1 D:121565 T G 0.4853
7 53932886 ARS‐BFGL‐NGS‐20197 5.71E − 07 PCDHB6 U:892 T C 0.4966
28 46248750 BovineHD2800013502 4.71E − 07 LOC100141022 D:32551 T C 0.2523
Rump angle 1 5519845 BTB‐00003652 1.76E − 06 GRIK1 Within C T 0.25
1 68909418 BovineHD0100019488 4.88E − 07 CCDC14 Within A G 0.4182
1 123851563 BovineHD0100034972 1.97E − 06     T C 0.2011
1 142967164 BovineHD0100041062 2.03E − 06 BACE2 Within G A 0.4354
2 127566752 BovineHD0200037025 6.11E − 07 PDIK1L Within G A 0.3417
4 10148342 Hapmap35652‐SCAFFOLD151622_1051 4.57E − 07 LOC100295705 D:88944 C T 0.2268
6 83512619 BTA‐107087‐no‐rs 2.18E − 06 LOC100298985 U:19994 G A 0.3243
6 87715723 Hapmap38371‐BTA‐105598 1.58E − 06 AMBN D:9991 C A 0.3356
7 83260664 BovineHD0700024393 4.32E − 09 MSH3 D:33418 G A 0.1746
7 83757564 BovineHD0700024587 1.14E − 07 SSBP2 Within C A 0.11
7 91507089 ARS‐BFGL‐NGS‐118534 2.83E − 07     G A 0.2761
8 101664818 BovineHD0800030195 2.25E − 06 SVEP1 Within G A 0.1236
9 81329823 BTA‐106078‐no‐rs 9.84E − 07 HIVEP2 D:62953 A G 0.4592
10 43438784 BovineHD1000013067 8.09E − 08 MAP4K5 Within G A 0.3246
10 62563388 BovineHD1000018043 7.73E − 07 SLC24A5 D:73734 T C 0.4376
10 73979984 Hapmap49737‐BTA‐75278 6.00E − 07 PRKCH D:52860 A G 0.04762
18 55309510 ARS‐BFGL‐NGS‐116541 2.37E − 06 LIG1 Within C A 0.4487
18 55514759 BovineHD1800016250 7.28E − 08 SYNGR4 Within T C 0.2494
18 55621823 ARS‐BFGL‐NGS‐31529 2.12E − 06 LMTK3 Within T G 0.2426
22 47989704 BovineHD2200013812 1.72E − 06 CACNA1D Within T C 0.2727
22 48408579 BovineHD2200013926 5.34E − 07 RFT1 Within G A 0.0839
24 29554807 BovineHD2400008037 2.70E − 07 LOC782418 D:63567 G A 0.2323
25 31147780 Hapmap24744‐BTC‐028427 1.82E − 08 LOC100301342 Within T C 0.3898
25 42364359 ARS‐BFGL‐NGS‐101981 1.32E − 06 ADAP1 D:1441 T C 0.2823
26 16504170 BovineHD2600004135 1.32E − 06 LOC522146 Within A G 0.1927
29 7216410 BovineHD2900002021 5.13E − 07 LOC100336861 Within G A 0.1961
30 2037499 BovineHD3000000680 2.28E − 06 KLHL13 Within C A 0.03855
30 141936249 BTA‐21001‐no‐rs 2.16E − 06 MSL3 U:105193 C T 0.1939
Final conformation score Final score 6 1770665 BovineHD0600000461 1.75E − 06 1‐Mar Within G A 0.39
11 80730546 ARS‐BFGL‐NGS‐41612 4.61E − 07 KCNS3 U:112015 G A 0.1485
13 28331553 ARS‐BFGL‐NGS‐109467 1.80E − 06 SEPHS1 D:16954 A G 0.4388
16 37787772 ARS‐BFGL‐NGS‐34764 2.01E − 06 NME7 Within G A 0.4887
16 37904090 BTB‐00637941 1.17E − 06 SLC19A2 Within C T 0.4863
16 41384258 BovineHD1600011634 2.18E − 06 LOC614226 U:123143 T C 0.4558
18 4463083 Hapmap57004‐rs29011610 1.17E − 06 MON1B D:5928 G A 0.3129
18 65405023 BovineHD1800019051 2.04E − 06 LOC789374 Within A G 0.1179
19 26049717 BovineHD1900007686 7.31E − 07 AIPL1 D:7711 T C 0.03628
21 57552028 BovineHD2100016535 8.46E − 07 SLC24A4 U:44433 A G 0.234
25 39558290 BovineHD2500011031 2.24E − 06 WIPI2 U:7420 T G 0.09524
25 40192570 BovineHD4100017518 1.07E − 06 SDK1 Within T C 0.03061
26 44324248 BovineHD2600012439 1.82E − 06 OAT U:53908 C A 0.3379
30 12103258 BovineHD3000003945 8.38E − 07 ACTRT1 U:55118 G A 0.1497
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1

Chromosome 30 refers to X chromosome.

2

The distance from the SNP locus to the gene (unit: bp); D and U indicate that the SNP site is located downstream and upstream of the gene, respectively; ‘Within’ indicates that the SNP locus is located within the gene.

Table 2.

DNA regions of chromosomes 18, 21, 25 and X containing SNPs that were significantly associated with body conformation traits of dairy cows

Chromosome Trait Position SNP name P‐Value Nearest gene Distance 1 Reference Alteration MAF
18 AW 65402237 BovineHD1800019049 4.75E − 07 LOC789374 Within C T 0.2143
FS 4463083 Hapmap57004‐rs29011610 1.17E − 06 MON1B D: 5928 G A 0.3129
FS 65405023 BovineHD1800019051 2.04E − 06 LOC789374 Within A G 0.1179
MSS 55956772 ARS‐BFGL‐NGS‐60829 6.86E − 07 NUCB1 D: 1160 C T 0.4172
RA 55309510 ARS‐BFGL‐NGS‐116541 2.37E − 06 LIG1 Within C A 0.4487
RA 55514759 BovineHD1800016250 7.28E − 08 SYNGR4 Within T C 0.2494
RA 55621823 ARS‐BFGL‐NGS‐31529 2.12E − 06 LMTK3 Within T G 0.2426
21 FS 57552028 BovineHD2100016535 8.46E − 07 SLC24A4 U: 44433 A G 0.2340
AW 57583470 BovineHD2100016545 2.26E − 06 SLC24A4 U: 12991 A G 0.4478
AW 57584406 BovineHD2100016546 2.33E − 06 SLC24A4 U: 12055 A C 0.4477
AW 57587712 BovineHD2100016549 2.26E − 06 SLC24A4 U: 8749 C T 0.4478
AW 57620878 BovineHD2100016561 9.13E − 07 SLC24A4 Within G A 0.2823
AW 57623572 BovineHD2100016563 9.13E − 07 SLC24A4 Within G A 0.2823
25 RA 42364359 ARS‐BFGL‐NGS‐101981 1.32E − 06 ADAP1 D: 1441 T C 0.2823
UD 35623801 ARS‐BFGL‐NGS‐28167 2.16E − 06 CUX1 D: 12155 A G 0.4023
UD 36266951 BovineHD2500010029 1.27E − 07 LOC100298352 Within T C 0.3560
RA 31147780 Hapmap24744‐BTC‐028427 1.82E − 08 LOC100301342 Within T C 0.3898
FS 40192570 BovineHD4100017518 1.07E − 06 SDK1 Within T C 0.03061
X FS 12103258 BovineHD3000003945 8.38E − 07 ACTRT1 U: 55118 G A 0.1497
RA 2037499 BovineHD3000000680 2.28E − 06 KLHL13 Within C A 0.03855
AG 116273044 BovineHD3000032546 6.55E − 07 LOC537655 Within C A 0.3061
MS 138304543 BovineHD3000039710 5.31E − 07 LOC782196 U: 18403 T C 0.2130
AG 110222328 BTA‐116883‐no‐rs 1.56E − 06 LOC786124 D: 27320 G A 0.2404
AG 132705219 BovineHD3000037672 5.10E − 08 LOC786725 U: 54284 T C 0.4490
MSS 132705219 BovineHD3000037672 7.03E − 08 LOC786725 U: 54284 T C 0.4490
RA 141936249 BTA‐21001‐no‐rs 2.16E − 06 MSL3 U: 105193 C T 0.1939
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AW, Attachment width; FS, conformation final score; MSS, mammary system score; RA, rump angle; UD, udder depth; AG, angularity; MS, median suspensory; MAF means minor allele frequency.

1

The distance from the SNP locus to the gene (unit: bp); D and U indicate that the SNP site is located downstream and upstream of the gene, respectively; ‘Within’ indicates that the SNP locus is located within the gene.

In summary, a GWAS using linear statistical model was conducted on 29 body conformation traits in a Chinese Holstein cattle population, and 79 SNPs were found to have genome‐wide‐significant (P < 0.05) association effects on 14 body conformation traits. Among these significant SNPs, 74 of them are newly detected in this study, five have been reported in previous literature and 26 are located in genes and are worth further investigation to potentially identify the causative mutations underlying the QTL.

Conflict of interest

The authors declare that there is no conflict of interest.

Supporting information

Figure S1 Genome‐wide plots of −log10 (P‐value) SNP association effects on body conformation traits of rump angle (RA, a) and final score (FS, b) obtained by mixed‐effect linear regression model.

Click here for additional data file. (245.7KB, pdf)

Table S1 Descriptive statistics of the 29 conformation traits used in the GWAS.

Table S2 Distribution of SNP markers by chromosomes before and after quality control.

Table S3 The chromosomal distribution of significant SNPs associated with body conformation traits.

Table S4 The SNPs identified associated with multiple body conformation traits of dairy cows.

Table S5 Multiple SNPs located in the SLC24A4 gene region were significantly associated with body conformation traits of dairy cows.

Click here for additional data file. (199.5KB, doc)

Acknowledgements

The authors would like to thank the owners of the four farms who generously allowed us to sample hair follicles from their cattle. We are also thankful to Dr Chunsheng Ma, Dr Jizhen Wang, Dr Sihao Wu and Dr Enfa Wang at Alta‐Agricorp Co. Ltd for their kind support and suggestions on the verification of phenotype data. We are grateful to Dr Changsong Qian at Neogen Bio‐Scientific Technology (Shanghai) Co. Ltd for his technical support during the hair follicle collection. This work was financially supported by the Key Project of Agricultural Fine Breeding of Shandong Province (2016LZGC030, 2019LZGC011), the National Natural Science Foundation of China (31402054), the Natural Science Foundation of Shandong (ZR2013CM013), Funds of the Shandong ‘Double Tops’ Program (SYL2017YSTD08), the Modern Agricultural Industry Technology System (CARS‐36) and the Tai Mountain Scholar Program.

References

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

Figure S1 Genome‐wide plots of −log10 (P‐value) SNP association effects on body conformation traits of rump angle (RA, a) and final score (FS, b) obtained by mixed‐effect linear regression model.

Click here for additional data file. (245.7KB, pdf)

Table S1 Descriptive statistics of the 29 conformation traits used in the GWAS.

Table S2 Distribution of SNP markers by chromosomes before and after quality control.

Table S3 The chromosomal distribution of significant SNPs associated with body conformation traits.

Table S4 The SNPs identified associated with multiple body conformation traits of dairy cows.

Table S5 Multiple SNPs located in the SLC24A4 gene region were significantly associated with body conformation traits of dairy cows.

Click here for additional data file. (199.5KB, doc)

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