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  • MicroRNA-302 switch to identify and eliminate undifferentiated human pluripotent stem cells.

MicroRNA-302 switch to identify and eliminate undifferentiated human pluripotent stem cells.

Scientific reports (2016-09-10)
Callum J C Parr, Shota Katayama, Kenji Miki, Yi Kuang, Yoshinori Yoshida, Asuka Morizane, Jun Takahashi, Shinya Yamanaka, Hirohide Saito
ABSTRACT

The efficiency of pluripotent stem cell differentiation is highly variable, often resulting in heterogeneous populations that contain undifferentiated cells. Here we developed a sensitive, target-specific, and general method for removing undesired cells before transplantation. MicroRNA-302a-5p (miR-302a) is highly and specifically expressed in human pluripotent stem cells and gradually decreases to basal levels during differentiation. We synthesized a new RNA tool, miR-switch, as a live-cell reporter mRNA for miR-302a activity that can specifically detect human induced pluripotent stem cells (hiPSCs) down to a spiked level of 0.05% of hiPSCs in a heterogeneous population and can prevent teratoma formation in an in vivo tumorigenicity assay. Automated and selective hiPSC-elimination was achieved by controlling puromycin resistance using the miR-302a switch. Our system uniquely provides sensitive detection of pluripotent stem cells and partially differentiated cells. In addition to its ability to eliminate undifferentiated cells, miR-302a switch also holds great potential in investigating the dynamics of differentiation and/or reprograming of live-cells based on intracellular information.

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Anti-Polysialic Acid-NCAM Antibody, clone 2-2B, ascites fluid, clone 2-2B, Chemicon®