Modification of alternative splicing in bovine somatic cell nuclear transfer embryos using engineered CRISPR-Cas13d
Modification of alternative splicing in bovine somatic cell nuclear transfer embryos using engineered CRISPR-Cas13d
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使用工程改造的 CRISPRCas13d 修饰牛体细胞核移植胚胎中的选择性剪接
DOI：
10.1007/s11427-021-2060-x
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发表时间：
2022-05-05
影响因子：
9.1
通讯作者：
Liu, Xu
中科院分区：
文献类型：
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作者：
Cheng, Rui;Zheng, Xiaoman;Liu, Xu
动物克隆可以通过体细胞核移植(SCNT)实现，但由此产生的活出生率相对较低。我们以前通过外源褪黑素处理或过表达赖氨酸特异性去甲基酶4D(KDM4D)和4E(KDM4E)来提高牛SCNT的效率。在这项研究中，我们揭示了在受精和胚胎基因组激活过程中大量的选择性剪接(AS)转换，并发现与体外受精胚胎相比，牛SCNT胚胎中的AS异常。我们使用CRISPR-Cas13d RNA打靶系统靶向ABI2和ZNF106前-mRNA的顺式元件来修饰AS，从而使SCNT胚胎的异型率降低了近50%，并获得了较高的存活率(11%-19%)。这些结果表明，该系统可能为牛的克隆提供了一种有效的方法，同时也为进一步提高SCNT的效率铺平了道路。
Animal cloning can be achieved by somatic cell nuclear transfer (SCNT), but the resulting live birth rate is relatively low. We previously improved the efficiency of bovine SCNT by exogenous melatonin treatment or by overexpression of lysine-specific demethylase 4D (KDM4D) and 4E (KDM4E). In this study, we revealed abundant alternative splicing (AS) transitions during fertilization and embryonic genome activation, and demonstrated abnormal AS in bovine SCNT embryos compared with in vitro fertilized embryos. We used the CRISPR-Cas13d RNA-targeting system to target cis-elements of ABI2 and ZNF106 pre-mRNA to modify AS, thus reducing the ratio of abnormal-isoform SCNT embryos by nearly 50% and achieving a high survival rate (11%-19%). These results indicate that this system may provide an efficient method for bovine cloning, while also paving the way for further improvements in the efficiency of SCNT.