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Introducing the Stem Cell ASCL2 Reporter STAR into Intestinal Organoids.

STAR protocols (2020-12-31)
Maria C Heinz, Koen C Oost, Hugo J G Snippert
ZUSAMMENFASSUNG

Patient-derived organoids maintain functional and phenotypic characteristics of the original tissue such as cell-type diversity. Here, we provide protocols on how to label intestinal (cancer) stem cells by integrating the stem cell ASCL2 reporter (STAR) into human and mouse genomes via two different strategies: (1) lentiviral transduction or (2) transposon-based integration. Organoid technology, in combination with the user-friendly nature of STAR, will facilitate basic research in human and mouse adult stem cell biology. For complete details on the use and execution of this protocol, please refer to Oost et al. (2018).

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Dulbecco Modifiziertes Eagle-Medium – hoher Glucosegehalt, With 4500 mg/L glucose, L-glutamine, sodium pyruvate, and sodium bicarbonate, liquid, sterile-filtered, suitable for cell culture
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N-Acetyl-L-Cystein, BioReagent, suitable for cell culture
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Nicotinamid, BioReagent, suitable for cell culture, suitable for insect cell culture
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Puromycin -dihydrochlorid aus Streptomyces alboniger, ≥98% (HPLC), powder
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Hexadimethrinbromid, ≥95%
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Trypsin-Inhibitor aus Glycine max (soybean), lyophilized powder