Transcription activator-like effector nuclease-mediated deletion safely eliminates the major egg allergen ovomucoid in chickens
Transcription activator-like effector nuclease-mediated deletion safely eliminates the major egg allergen ovomucoid in chickens
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转录激活剂样效应物核酸酶介导的删除安全地消除了鸡中主要的鸡蛋过敏原卵类粘蛋白
DOI：
10.1016/j.fct.2023.113703
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发表时间：
2023
影响因子：
4.3
通讯作者：
Horiuchi Hiroyuki
中科院分区：
文献类型：
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作者：
Ezaki Ryo;Sakuma Tetsushi;Kodama Daisuke;Sasahara Ryou;Shiraogawa Taichi;Ichikawa Kennosuke;Matsuzaki Mei;Handa Akihiro;Yamamoto Takashi;Horiuchi Hiroyuki
在主要的鸡蛋过敏原中，卵类粘蛋白（OVM）对热和消化酶非常稳定，使得其难以通过物理化学方法去除。然而，最近的基因组编辑技术使产生OVM敲除鸡蛋成为可能。为了将这种OVM敲除的鸡蛋用作食品，重要的是评估其作为食品的安全性。因此，在这项研究中，我们检查了是否存在突变蛋白表达，载体序列插入和脱靶效应，在用铂TALENs敲除OVM的鸡中。纯合子OVM敲除母鸡产下的蛋没有明显的异常，免疫印迹显示蛋白中既不含成熟的OVM，也不含OVM截短的变体。全基因组测序（WGS）结果表明，TALEN诱导OVM基因敲除鸡的脱靶效应主要集中在基因间区和内含子区。WGS信息证实，用于基因组编辑的质粒载体仅瞬时存在，并且不整合到编辑的鸡的基因组中。这些结果表明了安全性评价的重要性，并揭示了这种OVM敲除鸡产下的鸡蛋解决了食品和疫苗的过敏问题。
Among the major egg allergens, ovomucoid (OVM) is very stable against heat and digestive enzymes, making it difficult to remove physiochemically and inactivate allergens. However, recent genome editing technology has made it possible to generate OVM-knockout chicken eggs. To use this OVM-knockout chicken egg as food, it is important to evaluate its safety as food. Therefore, in this study, we examined the presence or absence of mutant protein expression, vector sequence insertion, and off-target effects in chickens knocked out with OVM by platinum TALENs. The eggs laid by homozygous OVM-knockout hens showed no evident abnormalities, and immunoblotting showed that the albumen contained neither the mature OVM nor the OVM truncated variant. Whole genome sequencing (WGS) revealed that the potential TALEN-induced off-target effects inOVM-knockout chickens were localized in the intergenic and intron regions. The WGS information confirmed that plasmid vectors used for genome editing were only transiently present and did not integrate into the genome of edited chickens. These results indicate the importance of safety evaluation and reveal that the eggs laid by this OVM knockout chicken solve the allergy problem in food and vaccines.