Safety evaluation of myostatin-edited Meishan pigs by whole genome resequencing analyses

 

https://doi.org/10.17221/7/2018-CJASCitation:Xie S., Qian L., Cai C., Jiang S., Xiao G., Gao T., Li X., Cui W. (2019): Safety evaluation of myostatin-edited Meishan pigs by whole genome resequencing analyses  . Czech J. Anim. Sci., 64: 291-299.
download PDF

Genome editing technology can make specifically target genomic modifications, resulting in site specific DNA insertion, deletion or replacement in the genome of an organism. We have recently produced genetically engineered (GE) Meishan pigs containing a ZFN-edited myostatin (MSTN) loss-of-function mutation that leads to a clear “double muscle” phenotype as observed for Belgian cattle. In this study, whole genome resequencing was used as an approach to evaluate the safety risk, if any, associated with the introduction of a ZFN-edited myostatin (MSTN) loss-of-function mutation in a local pig breed, the Meishan pigs. The results of resequencing analyses show that the effective data from pigs of wild-type group and MSTN-edited GE group is greater than 99%. The 1× coverage rate is > 98%, and the 4× coverage rate is > 96%. The genetic variation on each chromosome is close to 1. From this whole genome resequencing study, our results demonstrated that 99.7% of single nucleotide polymorphisms (SNPs) are the same in the same genetic variation from both wild-type group and MSTN-edited GE group, implying genomic sequence variations are highly similar between the two groups of pigs.

References:
(2012): An integrated map of genetic variation from 1,092 human genomes. Nature, 491, 56-65  https://doi.org/10.1038/nature11632
 
Carlson Daniel F, Lancto Cheryl A, Zang Bin, Kim Eui-Soo, Walton Mark, Oldeschulte David, Seabury Christopher, Sonstegard Tad S, Fahrenkrug Scott C (2016): Production of hornless dairy cattle from genome-edited cell lines. Nature Biotechnology, 34, 479-481  https://doi.org/10.1038/nbt.3560
 
Carroll Dana (2011): Genome Engineering With Zinc-Finger Nucleases. Genetics, 188, 773-782  https://doi.org/10.1534/genetics.111.131433
 
Choi Jung-Woo, Liao Xiaoping, Park Sairom, Jeon Heoyn-Jeong, Chung Won-Hyong, Stothard Paul, Park Yeon-Soo, Lee Jeong-Koo, Lee Kyung-Tai, Kim Sang-Hwan, Oh Jae-Don, Kim Namshin, Kim Tae-Hun, Lee Hak-Kyo, Lee Sung-Jin (2013): Massively parallel sequencing of Chikso (Korean brindle cattle) to discover genome-wide SNPs and InDels. Molecules and Cells, 36, 203-211  https://doi.org/10.1007/s10059-013-2347-0
 
Choi Jung-Woo, Liao Xiaoping, Stothard Paul, Chung Won-Hyong, Jeon Heoyn-Jeong, Miller Stephen P., Choi So-Young, Lee Jeong-Koo, Yang Bokyoung, Lee Kyung-Tai, Han Kwang-Jin, Kim Hyeong-Cheol, Jeong Dongkee, Oh Jae-Don, Kim Namshin, Kim Tae-Hun, Lee Hak-Kyo, Lee Sung-Jin, te Pas Marinus FW. (2014): Whole-Genome Analyses of Korean Native and Holstein Cattle Breeds by Massively Parallel Sequencing. PLoS ONE, 9, e101127-  https://doi.org/10.1371/journal.pone.0101127
 
Choi Jung-Woo, Chung Won-Hyong, Lee Kyung-Tai, Cho Eun-Seok, Lee Si-Woo, Choi Bong-Hwan, Lee Sang-Heon, Lim Wonjun, Lim Dajeong, Lee Yun-Gyeong, Hong Joon-Ki, Kim Doo-Wan, Jeon Hyeon-Jeong, Kim Jiwoong, Kim Namshin, Kim Tae-Hun (2015): Whole-genome resequencing analyses of five pig breeds, including Korean wild and native, and three European origin breeds. DNA Research, 22, 259-267  https://doi.org/10.1093/dnares/dsv011
 
Fan Wen-Lang, Ng Chen Siang, Chen Chih-Feng, Lu Mei-Yeh Jade, Chen Yu-Hsiang, Liu Chia-Jung, Wu Siao-Man, Chen Chih-Kuan, Chen Jiun-Jie, Mao Chi-Tang, Lai Yu-Ting, Lo Wen-Sui, Chang Wei-Hua, Li Wen-Hsiung (2013): Genome-Wide Patterns of Genetic Variation in Two Domestic Chickens. Genome Biology and Evolution, 5, 1376-1392  https://doi.org/10.1093/gbe/evt097
 
Fang M., Hu X., Jiang T., Braunschweig M., Hu L., Du Z., Feng J., Zhang Q., Wu C., Li N. (2005): The phylogeny of Chinese indigenous pig breeds inferred from microsatellite markers. Animal Genetics, 36, 7-13  https://doi.org/10.1111/j.1365-2052.2004.01234.x
 
Gorodkin Jan, Cirera Susanna, Hedegaard Jakob, Gilchrist Michael J, Panitz Frank, Jørgensen Claus, Scheibye-Knudsen Karsten, Arvin Troels, Lumholdt Steen, Sawera Milena, Green Trine, Nielsen Bente J, Havgaard Jakob H, Rosenkilde Carina, Wang Jun, Li Heng, Li Ruiqiang, Liu Bin, Hu Songnian, Dong Wei, Li Wei, Yu Jun, Wang Jian, Stærfeldt Hans-Henrik, Wernersson Rasmus, Madsen Lone B, Thomsen Bo, Hornshøj Henrik, Bujie Zhan, Wang Xuegang, Wang Xuefei, Bolund Lars, Brunak Søren, Yang Huanming, Bendixen Christian, Fredholm Merete (): Porcine transcriptome analysis based on 97 non-normalized cDNA libraries and assembly of 1,021,891 expressed sequence tags. Genome Biology, 8, R45-  https://doi.org/10.1186/gb-2007-8-4-r45
 
Grobet L., Martin L.J., Poncelet D., Pirottin D., Brouwers B. (1997): A deletion in the bovine myostatin gene causes the double-muscled phenotype in cattle. Genome Biology, 17, 71–74.
 
Jones Huw D. (2015): Regulatory uncertainty over genome editing. Nature Plants, 1, -  https://doi.org/10.1038/nplants.2014.11
 
Kanis E., De Greef K. H., Hiemstra A., van Arendonk J. A. M. (2005): Breeding for societally important traits in pigs1. Journal of Animal Science, 83, 948-957  https://doi.org/10.2527/2005.834948x
 
Kerstens Hindrik HD, Kollers Sonja, Kommadath Arun, del Rosario Marisol, Dibbits Bert, Kinders Sylvia M, Crooijmans Richard P, Groenen Martien AM (2009): Mining for single nucleotide polymorphisms in pig genome sequence data. BMC Genomics, 10, 4-  https://doi.org/10.1186/1471-2164-10-4
 
Laible Götz, Wei Jingwei, Wagner Stefan (2015): Improving livestock for agriculture - technological progress from random transgenesis to precision genome editing heralds a new era. Biotechnology Journal, 10, 109-120  https://doi.org/10.1002/biot.201400193
 
Ledford Heidi (2016): Gene-editing surges as US rethinks regulations. Nature, 532, 158-159  https://doi.org/10.1038/532158a
 
Legault C. (1985): Selection of breeds, strains and individual pigs for prolificacy. Journal of Reproduction and Fertility Supplement, 33, 151–166.
 
Li H., Durbin R. (2009): Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics, 25, 1754-1760  https://doi.org/10.1093/bioinformatics/btp324
 
Li H., Handsaker B., Wysoker A., Fennell T., Ruan J., Homer N., Marth G., Abecasis G., Durbin R. (2009): The Sequence Alignment/Map format and SAMtools. Bioinformatics, 25, 2078-2079  https://doi.org/10.1093/bioinformatics/btp352
 
Lunney Joan K. (): Advances in Swine Biomedical Model Genomics. International Journal of Biological Sciences, , 179-184  https://doi.org/10.7150/ijbs.3.179
 
Luo Junjie, Song Zhiyuan, Yu Shengli, Cui Dan, Wang Benli, Ding Fangrong, Li Song, Dai Yunping, Li Ning, Isalan Mark (2014): Efficient Generation of Myostatin (MSTN) Biallelic Mutations in Cattle Using Zinc Finger Nucleases. PLoS ONE, 9, e95225-  https://doi.org/10.1371/journal.pone.0095225
 
Mosher Dana S., Quignon Pascale, Bustamante Carlos D., Sutter Nathan B., Mellersh Cathryn S., Parker Heidi G., Ostrander Elaine A. (2007): A Mutation in the Myostatin Gene Increases Muscle Mass and Enhances Racing Performance in Heterozygote Dogs. PLoS Genetics, 3, e79-  https://doi.org/10.1371/journal.pgen.0030079
 
Qian Lili, Tang Maoxue, Yang Jinzeng, Wang Qingqing, Cai Chunbo, Jiang Shengwang, Li Hegang, Jiang Ke, Gao Pengfei, Ma Dezun, Chen Yaoxing, An Xiaorong, Li Kui, Cui Wentao (2015): Targeted mutations in myostatin by zinc-finger nucleases result in double-muscled phenotype in Meishan pigs. Scientific Reports, 5, -  https://doi.org/10.1038/srep14435
 
Rehfeldt C., Fiedler I., Dietl G., Ender K. (2000): Myogenesis and postnatal skeletal muscle cell growth as influenced by selection. Livestock Production Science, 66, 177-188  https://doi.org/10.1016/S0301-6226(00)00225-6
 
Schuelke Markus, Wagner Kathryn R., Stolz Leslie E., Hübner Christoph, Riebel Thomas, Kömen Wolfgang, Braun Thomas, Tobin James F., Lee Se-Jin (2004): Myostatin Mutation Associated with Gross Muscle Hypertrophy in a Child. New England Journal of Medicine, 350, 2682-2688  https://doi.org/10.1056/NEJMoa040933
 
Stothard Paul, Choi Jung-Woo, Basu Urmila, Sumner-Thomson Jennifer M, Meng Yan, Liao Xiaoping, Moore Stephen S (2011): Whole genome resequencing of black Angus and Holstein cattle for SNP and CNV discovery. BMC Genomics, 12, -  https://doi.org/10.1186/1471-2164-12-559
 
Walters Eric M, Wolf Eckhard, Whyte Jeffery J, Mao Jiude, Renner Simone, Nagashima Hiroshi, Kobayashi Eiji, Zhao Jianguo, Wells Kevin D, Critser John K, Riley Lela K, Prather Randall S (2012): Completion of the swine genome will simplify the production of swine as a large animal biomedical model. BMC Medical Genomics, 5, -  https://doi.org/10.1186/1755-8794-5-55
 
Wang K., Li M., Hakonarson H. (2010): ANNOVAR: functional annotation of genetic variants from high-throughput sequencing data. Nucleic Acids Research, 38, e164-e164  https://doi.org/10.1093/nar/gkq603
 
Xiao Gao-jun, Jiang Sheng-Wang, Qian Li-Li, Cai Chun-Bo, Wang Qing-qing, Ma De-Zun, Li Biao, Xie Shan-shan, Cui Wen-Tao, Li Kui, Bader Michael (2016): A 90-Day Feeding Study in Rats to Assess the Safety of Genetically Engineered Pork. PLOS ONE, 11, e0165843-  https://doi.org/10.1371/journal.pone.0165843
 
download PDF

© 2019 Czech Academy of Agricultural Sciences