title: Quantitative Single-Cell mRNA Analysis in Hydrogel Beads.
abstract: In recent years, technologies capable of analyzing single cells have emerged that are transforming many fields of biological research. Herein we report how DNA-functionalized hydrogel beads can serve as a matrix to capture mRNA from lysed single cells. mRNA quantification free of pre-amplification bias is ensured by using padlock probes and rolling circle amplification followed by hybridization with fluorescent probes. The number of transcripts in individual cells is assessed by simply counting fluorescent dots inside gel beads. The method extends the potential of existing techniques and provides a general platform for capturing molecules of interest from single cells.
authors.affiliation: Institute for Molecules and Materials, Radboud University Nijmegen, Heyendaalseweg 135, 6525, AJ, Nijmegen, The Netherlands.
authors.forename: Agata
authors.initials: A
authors.lastname: Rakszewska
authors.affiliation: Institute for Molecules and Materials, Radboud University Nijmegen, Heyendaalseweg 135, 6525, AJ, Nijmegen, The Netherlands.
authors.forename: Rosa J
authors.initials: RJ
authors.lastname: Stolper
authors.affiliation: Warsaw University of Life Sciences-SGGW, Nowoursynowska 166 ST, 02-787, Warszawa, Poland.
authors.forename: Anna B
authors.initials: AB
authors.lastname: Kolasa
authors.affiliation: Institute for Molecules and Materials, Radboud University Nijmegen, Heyendaalseweg 135, 6525, AJ, Nijmegen, The Netherlands.
authors.forename: Aigars
authors.initials: A
authors.lastname: Piruska
authors.affiliation: Institute for Molecules and Materials, Radboud University Nijmegen, Heyendaalseweg 135, 6525, AJ, Nijmegen, The Netherlands. w.huck@science.ru.nl.
authors.forename: Wilhelm T S
authors.initials: WT
authors.lastname: Huck
mesh_terms: D000199:Actins / Q000235:genetics; D017931:DNA Primers / Q000737:chemistry / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D020100:Hydrogels / Q000737:chemistry*; D020014:K562 Cells; D021141:Nucleic Acid Amplification Techniques; D009693:Nucleic Acid Hybridization; D009841:Oligonucleotides / Q000737:chemistry; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis*; D059010:Single-Cell Analysis
chemical_list: D000199:Actins; D017931:DNA Primers; D005456:Fluorescent Dyes; D020100:Hydrogels; D009841:Oligonucleotides; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; C477371:locked nucleic acid
keywords: droplet microfluidics; gels; mRNA; rolling circle amplification; single-cell analysis