Genome Whole-genome sequencing association analysis reveals the genetic architecture of meat quality traits in Chinese Qingyu pigs Journal: Genome Manuscript ID gen-2019-0227.R3 Manuscript Type: Article Date Submitted by the 22-May-2020 Author: Complete List of Authors: Wu, Pingxian; Sichuan Agricultural University - Chengdu Campus Wang, Kai; Sichuan Agricultural University Zhou, Jie; Sichuan Agricultural University Chen, Dejuan;Draft Sichuan Agricultural University - Chengdu Campus Yang, Xidi ; Sichuan Agricultural University - Chengdu Campus Jiang, Anan; Sichuan Agricultural University - Chengdu Campus Shen, Linyuan ; Sichuan Agricultural University - Chengdu Campus Zhang, Shunhua ; Sichuan Agricultural University - Chengdu Campus Xiao, Weihang ; Sichuan Agricultural University - Chengdu Campus Jiang, Yanzhi; Sichuan Agricultural University Zhu, Li; Sichuan Agricultural University - Chengdu Campus Li, Xuewei ; Sichuan Agricultural University - Chengdu Campus Xu, Xu; Sichuan Provincial Animal Husbandry and Food Bureau Zeng, Yangshuang; Sichuan Provincial Animal Husbandry and Food Bureau Tang, Guoqing ; Sichuan Agricultural University - Chengdu Campus Keyword: Pork color, Pork pH, GWAS, SNPs, Qingyu pigs Is the invited manuscript for consideration in a Special Not applicable (regular submission) Issue? : https://mc06.manuscriptcentral.com/genome-pubs Page 1 of 42 Genome 1 Title Page 2 Article Title 3 Whole-genome sequencing association analysis reveals the genetic architecture of 4 meat quality traits in Chinese Qingyu pigs 5 Authors 6 Pingxian Wu 1,†, Kai Wang 1,†, Jie Zhou 1, Dejuan Chen 1 , Xidi Yang 1, Anan Jiang 1, 7 Linyuan Shen1, Shunhua Zhang 1, Weihang Xiao 1, Yanzhi Jiang 2, Li Zhu 1 , Yangshuang 8 Zeng3, Xu Xu3, Xuewei Li 1, Guoqing Tang 1,* 9 1 Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of 10 Sichuan Province, Sichuan Agricultural University, Chengdu 611130, Sichuan, China 11 2 College of Life Science, Sichuan Agricultural University, Yaan 625014, Sichuan, China 12 3 Sichuan Animal Husbandry Station, Chengdu, 610041, Sichuan, China 13 * Corresponding author : GuoqingDraft Tang, [email protected]. 14 † Co-first author : These authors contributed equally to this work. 15 Authors’ Email 16 Pingxian Wu, Email: [email protected]. 17 Kai Wang, Email: [email protected]. 18 Jie Zhou, Email: [email protected]. 19 Dejuan Chen, Email: [email protected]. 20 Xidi Yang, Email: [email protected]. 21 Shunhua Zhang, Email: [email protected]. 22 Anan Jiang, Email: [email protected]. 23 Linyuan Shen, Email: [email protected]. 24 Weihang Xiao, Email: [email protected]. 25 Yanzhi Jiang, Email: [email protected]. 26 Li Zhu, Email: [email protected]. 27 Xuewei Li, Email: [email protected]. 28 Yangshuang Zeng, Email: [email protected]. 29 Xu Xu, Email: [email protected]. 30 Guoqing Tang, Email: [email protected]. https://mc06.manuscriptcentral.com/genome-pubs Genome Page 2 of 42 31 Whole-genome sequencing association analysis reveals the 32 genetic architecture of meat quality traits in Chinese Qingyu 33 pigs 34 Abstract 35 The Chinese Qingyu pig breed is an invaluable indigenous genetic resource. 36 However, few studies have investigated the genetic architecture of meat 37 quality traits in Qingyu pigs. Here, 30 purebred Qingyu pigs were 38 subjected to whole-genome sequencing. After quality control, 18,436,759 39 SNPs were retained. Genome-wide association studies (GWAS) were then 40 performed for meat pH and Draftcolor at three postmortem time points (45 41 min, 24 h and 48 h) using single-marker regression analysis. In total, 11 42 and 69 SNPs were associated with meat pH and color of the longissimus 43 thoracis muscle (LTM), respectively, while 54 and 29 SNPs were associated 44 with meat pH and color of the semimembranosus muscle (SM), 45 respectively. Seven SNPs associated with pork pH were shared by all three 46 postmortem time points. Several candidate genes for meat traits were 47 identified, including four genes (CXXC5, RYR3, BNIP3, and MYCT1) related 48 to skeletal muscle development, regulation of Ca2+ release in the muscle, 49 and anaerobic respiration, which are promising candidates for selecting 50 superior meat quality traits in Qingyu pigs. To our knowledge, this is the 51 first study investigating the postmortem genetic architecture of pork pH 52 and color in Qingyu pigs. Our findings further the current understanding https://mc06.manuscriptcentral.com/genome-pubs Page 3 of 42 Genome 53 of the genetic factors influencing meat quality. 54 Keywords: Pork color, Pork pH, GWAS, SNPs, Qingyu pigs 55 56 Introduction 57 The Chinese Qingyu pig breed is an invaluable Chinese genetic resource 58 (China National Commission of Animal Genetic Resources, 2012). Qingyu 59 pigs are mainly distributed in a narrow region within the mountainous 60 areas of Sichuan Province, China. It is one of the most famous indigenous 61 pig breeds for its tender meat and tasty flavor(Linyuan Shen, 2014). 62 Previously, these pigs wereDraft mainly subjected to long-term natural 63 selection and short-term traditional selection programs. Thus, their 64 genome is likely markedly different from that of other pig breeds. Recent 65 investigations have focused on meat quality traits in pigs(Verardo, Sevón- 66 Aimonen, Serenius, Hietakangas, & Uimari, 2017; Zhou et al., 2014), 67 however, because of their limited population size, no similar study has 68 been performed on Qingyu pigs at whole-genome level. 69 Meat quality is one of the most important economic factors in pig 70 production. However, because meat quality is measured after slaughter, 71 it is difficult and expensive to obtain the phenotypes of meat quality. 72 Previous studies have reported that many pig meat qualities exhibit low 73 to moderate heritability, such as meat color (0.1 < h2 < 0.4), pH (h2 = 0.15), 74 shear force (h2 = 0.40), and drip loss (h2 = 0.20) in pigs(Miar et al., 2014). https://mc06.manuscriptcentral.com/genome-pubs Genome Page 4 of 42 75 More recently, several studies have been performed to identify the 76 candidate genes and genetic variants influencing meat quality 77 traits(Schmid, Maushammer, Preuß, & Bennewitz, 2018; Vidal, Noguera, 78 Amills, Varona, & Sánchez, 2005). A total of 16,071 quantitative trait loci 79 (QTLs) (PigQTLdb, https://www.animalgenome.org/cgi- 80 bin/QTLdb/SS/index, 2019) associated with meat and carcass traits in pigs. 81 Among these QTLs, 722 and 632 were found to be associated with meat 82 pH and color, respectively. However, most of these QTLs had large 83 confidence intervals and lacked reproducibility in different populations. 84 To date, no QTLs have been Draftidentified for complex traits in Qingyu pigs 85 specifically. However, identifying variants in key genomic regions is 86 important for genetic improvement of this breed, because traditional 87 selection methods do not allow selection of highly specific meat quality 88 traits. 89 Genome-wide association studies (GWAS) provide a systematic and 90 powerful approach for gaining deeper insights into complex traits in 91 livestock. Using SNP arrays, GWAS for meat quality traits have identified 92 many QTLs and candidate genes in commercial populations(Casiró et al., 93 2017; Nonneman et al., 2013; Sanchez, Tribout, Iannuccelli, Bouffaud, 94 Servin, Tenghe, Dehais, Muller, Del Schneider, et al., 2014; Uimari, Sironen, 95 & Sevón-Aimonen, 2013; C. Zhang et al., 2015). However, SNP arrays are 96 limited in that they only identify a small fraction of genetic variants, https://mc06.manuscriptcentral.com/genome-pubs Page 5 of 42 Genome 97 resulting in low power of the GWAS, whereas whole-genome sequencing 98 (WGS) can potentially detect all genetic variants. Using WGS data, the 99 accuracy and power of GWAS are significantly improved, allowing credible 100 associations to be obtained(Daetwyler et al., 2014). Such advances make 101 genetic improvement achievable for meat quality traits in pigs. 102 Previous studies identified several candidate genes (such as PRKAG3, 103 RYR1 and IGF2 gene) affecting meat quality traits in lean-type western 104 pigs, and these findings facilitated considerable improvements in meat 105 quality(Gao, Zhang, Hu, & Li, 2007; Vries, Faucitano, Sosnicki, & Plastow, 106 2000). However, the Chinese DraftQingyu pig breed is a fat-type indigenous pig 107 line, and there are obvious differences between common commercial pigs 108 and Qingyu pigs in meat quality and productivity. Consequently, it is likely 109 that these breeds would exhibit significant genetic differences. 110 In this study, we performed a GWAS on meat pH and color at three 111 different times after slaughter (45 min, 24 h, and 48 h postmortem), using 112 WGS data from a purebred Qingyu pig population. The objective was to 113 identify candidate genes and genetic variants associated with meat pH 114 and color in Qingyu pigs. 115 Materials and methods 116 Ethics statement 117 All animals procedures related to this study were approved by the 118 Institutional Review Board (IRB14044) and the Institutional Animal Care https://mc06.manuscriptcentral.com/genome-pubs Genome Page 6 of 42 119 and Use Committee of the Sichuan Agricultural University under permit 120 number DKY–B20140302. 121 Animals and phenotypes 122 A total of 30 purebred Qingyu pigs (including 15 females and 15 males) 123 from the Bashan animal husbandry technology Co., LTD. were used in this 124 study. During the period of 2018-2019, all pigs were subjected to same 125 feeding conditions. These pigs