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Measuring MicroRNA Expression in Mouse Hematopoietic Stem Cells

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Hematopoietic Stem Cell Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1185))

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Abstract

MicroRNAs (miRNAs) are important regulators of diverse biologic processes. In the hematopoietic system, miRNAs have been shown to regulate lineage fate decisions, mature immune effector cell function, apoptosis, and cell cycling, and a more limited number of miRNAs has been shown to regulate hematopoietic stem cell (HSC) self-renewal. Many of these miRNAs were initially identified as candidate regulators of HSC function by comparing miRNA expression in hematopoietic stem and progenitors cells (HSPCs) to their mature progeny. While the measurement of miRNA expression in rare cell populations such as HSCs poses practical challenges due to the low amount of RNA present, a number of techniques have been developed to measure miRNAs in small numbers of cells. Here, we describe our protocol for measuring miRNAs in purified mouse HSCs using a highly sensitive real-time quantitative PCR strategy that utilizes microfluidic array cards containing pre-spotted TaqMan probes that allows the detection of mature miRNAs in small reaction volumes. We also describe a simple data analysis method to evaluate miRNA expression profiling data using an open-source software package (HTqPCR) using mouse HSC miRNA profiling data generated in our lab.

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Acknowledgments

This work was supported by a New Investigator Award in Bone Marrow Failure from the Department of Defense as well as an R01 award (1R01CA164120-01A1) from the NIH/NCI (to CYP).

Author information

Authors and Affiliations

  1. Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA

    Wenhuo Hu

  2. Human Oncology and Pathogenesis Program, Departments of Pathology and Laboratory Medicine, Memorial Sloan Kettering Cancer Center, 1275 York Ave., Box 20, New York, NY, 10065, USA

    Christopher Y. Park

Authors
  1. Wenhuo Hu
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  2. Christopher Y. Park
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Corresponding author

Correspondence to Christopher Y. Park .

Editor information

Editors and Affiliations

  1. Department of Pediatrics, Aflac Cancer and Blood Disorders Center Emory University School of Medicine, Atlanta, Georgia, USA

    Kevin D. Bunting

  2. Department of Pediatrics, Emory University School of Medicine Aflac Cancer & Blood Disorders Ctr, Atlanta, Georgia, USA

    Cheng-Kui Qu

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Hu, W., Park, C.Y. (2014). Measuring MicroRNA Expression in Mouse Hematopoietic Stem Cells. In: Bunting, K., Qu, CK. (eds) Hematopoietic Stem Cell Protocols. Methods in Molecular Biology, vol 1185. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1133-2_8

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  • DOI: https://doi.org/10.1007/978-1-4939-1133-2_8

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1132-5

  • Online ISBN: 978-1-4939-1133-2

  • eBook Packages: Springer Protocols

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