||Generation of Mouse Haploid Somatic Cells by Small Molecules for Genome-wide Genetic Screening|
||Zheng-Quan He5, Bao-Long Xia5, Yu-Kai Wang5, Jing Li, Gui-Hai Feng, Lin-Lin Zhang, Yu-Huan Li, Hai-Feng Wan, Tian-Da Li, Kai Xu, Xue-Wei Yuan, Yu-Fei Li, Xin-Xin Zhang, Ying Zhang, Liu Wang, Wei Li, Qi Zhou|
The recent success of derivation of mammalian haploid embryonic stem cells (haESCs) has provided a powerful tool for large-scale functional analysis of the mammalian genome. However, haESCs rapidly become diploidized after differentiation, posing challenges for genetic analysis. Here, we show that the spontaneous diploidization of haESCs happens in metaphase due to mitotic slippage. Diploidization can be suppressed by small-molecule-mediated inhibition of CDK1 and ROCK. Through ROCK inhibition, we can generate haploid somatic cells of all three germ layers from haESCs, including terminally differentiated neurons. Using piggyBac transposon-based insertional mutagenesis, we generated a haploid neural cell library harboring genome-wide mutations for genetic screening. As a proof of concept, we screened for Mn2+-mediated toxicity and identified the Park2 gene. Our findings expand the applications of mouse haploid cell technology to somatic cell types and may also shed light on the mechanisms of ploidy maintenance.
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