PuSH - Publication Server of Helmholtz Zentrum München

Export: TXT Text RIS Endnote (RIS) BIB BibTeX
Lahm, H.* ; Doppler, S.A.* ; Dressen, M.* ; Werner, A.* ; Adamczyk, K.* ; Schrambke, D.* ; Brade, T.* ; Laugwitz, K.L.* ; Deutsch, M.A.* ; Schiemann, M. ; Lange, R.* ; Moretti, A.* ; Krane, M.*

Live fluorescent RNA-based detection of pluripotency gene expression in embryonic and induced pluripotent stem cells of different species.

Stem Cells 33, 392-402 (2015)
Publ. Version/Full Text DOI
Free by publisher
The generation of induced pluripotent stem (iPS) cells has successfully been achieved in many species. However, the identification of truly reprogrammed iPS cells still remains laborious and the detection of pluripotency markers requires fixation of cells in most cases. Here, we report an approach with nanoparticles carrying Cy3-labeled sense oligonucleotide reporter strands coupled to gold-particles. These molecules are directly added to cultured cells without any manipulation and gene expression is evaluated microscopically after overnight incubation. To simultaneously detect gene expression in different species, probe sequences were chosen according to interspecies homology. With a common target-specific probe we could successfully demonstrate expression of the GAPDH house-keeping gene in somatic cells and expression of the pluripotency markers NANOG and GDF3 in embryonic stem cells and iPS cells of murine, human, and porcine origin. The population of target gene positive cells could be purified by fluorescence-activated cell sorting. After lentiviral transduction of murine tail-tip fibroblasts Nanog-specific probes identified truly reprogrammed murine iPS cells in situ during development based on their Cy3-fluorescence. The intensity of Nanog-specific fluorescence correlated positively with an increased capacity of individual clones to differentiate into cells of all three germ layers. Our approach offers a universal tool to detect intracellular gene expression directly in live cells of any desired origin without the need for manipulation, thus allowing conservation of the genetic background of the target cell. Furthermore, it represents an easy, scalable method for efficient screening of pluripotency which is highly desirable during high-throughput cell reprogramming and after genomic editing of pluripotent stem cells.
Impact Factor
Scopus SNIP
Web of Science
Times Cited
Scopus
Cited By
Altmetric
7.133
1.554
26
24
Tags
Annotations
Special Publikation
Hide on homepage

Edit extra information
Edit own tags
Private
Edit own annotation
Private
Hide on publication lists
on hompage
Mark as special
publikation
Publication type Article: Journal article
Document type Scientific Article
Keywords Fluorescent Nanoparticles ; Live Staining ; Pluripotency ; Embryonic Stem Cells ; Induced Pluripotent Stem Cells; Long-qt Syndrome; Defined Factors; Smooth-muscle; Ips Cells; In-vitro; Generation; Lines; Fibroblasts; Cardiomyocytes; Lineages
Language english
Publication Year 2015
HGF-reported in Year 2015
ISSN (print) / ISBN 0737-1454
e-ISSN 1549-4918
Journal Stem Cells
Quellenangaben Volume: 33, Issue: 2, Pages: 392-402 Article Number: , Supplement: ,
Publisher Wiley
Publishing Place Hoboken
Reviewing status Peer reviewed
Institute(s) Institute of Molecular Immunology (IMI)
POF-Topic(s) 30504 - Mechanisms of Genetic and Environmental Influences on Health and Disease
Research field(s) Immune Response and Infection
PSP Element(s) G-501790-003
DOI 10.1002/stem.1872
WOS ID WOS:000348211300009
Erfassungsdatum 2015-02-23
[➜Report correction]