title: Next Generation RNA Sequencing Analysis Reveals Expression of a Transient EMT Profile During Early Organization of MCF10A Cells in 3D.
abstract: RNA sequencing is a technique widely used to identify and characterize gene expression patterns. We demonstrate that this method can be applied to screen expression profiles in mammary epithelial cells cultured in 3D, supported by a natural laminin-rich extracellular matrix, but requires several specific steps in the preparation of the RNA samples. Here we describe the use of RNA sequencing to analyze mRNA patterns in MCF10A human mammary epithelial cells cultured under 3D conditions in a laminin-rich extracellular matrix. We focus on our methods for total RNA extraction at early time points during the formation and maturation of 3D acinus structures in these cultures and provide examples of our results and downstream analysis.
authors: Systems Biology Ireland, University College Dublin, Belfield, Dublin, 4, Ireland.|UCD School of Biomedical and Biological Science, University College Dublin, Belfield, Dublin, 4, Ireland.
authors: Benedikt
authors: B
authors: Minke
authors: Systems Biology Ireland, University College Dublin, Belfield, Dublin, 4, Ireland.
authors: Drieke
authors: D
authors: Vandamme
authors: Systems Biology Ireland, University College Dublin, Belfield, Dublin, 4, Ireland.
authors: Thomas
authors: T
authors: Schwarzl
authors: Systems Biology Ireland, University College Dublin, Belfield, Dublin, 4, Ireland.|UCD Conway Institute, University College Dublin, Belfield, Dublin, 4, Ireland.
authors: Walter
authors: W
authors: Kolch
authors: UCD School of Biomolecular and Biomedical Science, UCD Conway Institute, University College Dublin, Belfield, Dublin, Ireland. finian.martin@ucd.ie.
authors: Finian
authors: F
authors: Martin
mesh_terms: D061354:Acinar Cells / Q000378:metabolism; D045744:Cell Line, Tumor; D004847:Epithelial Cells / Q000378:metabolism; D058750:Epithelial-Mesenchymal Transition / Q000235:genetics*; D005109:Extracellular Matrix / Q000378:metabolism; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D006801:Humans; D007797:Laminin; D012333:RNA, Messenger / Q000235:genetics*; D017423:Sequence Analysis, RNA / Q000379:methods
chemical_list: D007797:Laminin; D012333:RNA, Messenger
keywords: Acinus; Acinus formation; EMT; Epithelial–mesenchymal transition; Id-1; MCF10A cells; Mammary epithelial cells; Next generation sequencing; RNA sequencing