Genetic Engineering of Human Embryonic Stem Cells for Precise Cell Fate Tracing during Human Lineage Development

Publication date: Available online 25 October 2018Source: Stem Cell ReportsAuthor(s): Zhenyu Chen, Xudong Ren, Xiangjie Xu, Xiaojie Zhang, Yi Hui, Zhongliang Liu, Lei Shi, Yujiang Fang, Lin Ma, Yang Liu, Daniel Terheyden-Keighley, Ling Liu, Xiaoqing ZhangSummaryIt is highly desirable to specify human developmental principles in an appropriate human model with advanced genetic tools. However, genetically engineering human cells with lineage-tracing systems has not been achieved. Here we introduce strategies to construct lineage-tracing systems in human embryonic stem cells (hESCs). The AAVS1 locus was suitable for the integration of the conditional reporter. The Cre-LoxP and Flp-FRT systems were highly sensitive, which may cause inaccurate lineage labeling in human cells. The recombination sensitivity and tracing fidelity could be finely tuned by modification of the LoxP recombination site. Moreover, tamoxifen-controllable CreERT2-LoxP and FlpERT2-FRT systems showed compelling advantages in tightly tracing human lineages temporally. In proof-of-principle experiments, we traced human PAX6+ neuroectoderm cells and revealed their full neural lineage differentiation potency both in vitro and in vivo. Devising and optimizing of lineage-tracing systems in hESCs will thus set up a solid foundation for human developmental studies.Graphical Abstract
Source: Stem Cell Reports - Category: Stem Cells Source Type: research