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AssigneeGeron Corporation
InventorsWalter D. Funk, Melissa K. Carpenter, Joseph D. Gold, Margaret S. Inokuma, Chunhui Xu
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This disclosure provides a system for obtaining expression libraries from primate pluripotent stem (pPS) cells. pPS cells can be maintained in vitro without requiring a layer of feeder cells to inhibit differentiation. The role of the feeder cells is replaced by several other culture conditions provided in a suitable combination. Conditions that promote pPS cell growth without differentiation include supporting the culture on an extracellular matrix, and culturing the cells in a medium conditioned by another cell type. The cDNA libraries from such cultures are devoid of transcripts of feeder cell origin, relatively uncontaminated by transcripts from differentiated cells, and can have a high proportion of full-length transcripts. Subtraction libraries can also be produced that are enriched for transcripts modulated during differentiation.
A method of obtaining an mRNA preparation or a cDNA library from undifferentiated primate pluripotent stem (pPS) cells or their differentiated progeny, comprising:
A cDNA library of undifferentiated primate pluripotent stem (pPS) cells, which can be obtained by providing undifferentiated pPS cells cultured in a suitable nutrient medium on an extracellular matrix but essentially free of feeder cells, and isolating mRNA from the undifferentiated cells.
A cDNA library comprising cDNA from at least 1,000 genes expressed at the mRNA level, wherein the library has been obtained from undifferentiated pPS cells, and is essentially free of cDNA of other vertebrates.
A cDNA library of at least 1,000 genes in which at least 50% of the cDNA is from genes that are modulated during differentiation of pPS cells, essentially free of cDNA of other vertebrates.
Dependencies are inferred from the claim text.