Introducing the Stem Cell ASCL2 Reporter STAR into Intestinal Organoids.

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

ABSTRACT

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).

MATERIALS

Product Number

Brand

Product Description

107689

Sigma-Aldrich

Hexadimethrine bromide, ≥95%

P7255

Sigma-Aldrich

Puromycin dihydrochloride from Streptomyces alboniger, ≥98% (HPLC), powder

T9003

Sigma-Aldrich

Trypsin inhibitor from Glycine max (soybean), lyophilized powder

D6429

Sigma-Aldrich

Dulbecco′s Modified Eagle′s Medium - high glucose, With 4500 mg/L glucose, L-glutamine, sodium pyruvate, and sodium bicarbonate, liquid, sterile-filtered, suitable for cell culture

A9165

Sigma-Aldrich

N-Acetyl-L-cysteine, BioReagent, suitable for cell culture

N0636

Sigma-Aldrich

Nicotinamide, BioReagent, suitable for cell culture, suitable for insect cell culture