title: Localization of mRNA in vertebrate axonal compartments by in situ hybridization.
abstract: The conclusive demonstration of RNA in vertebrate axons by in situ hybridization (ISH) has been elusive. We review the most important reasons for difficulties, including low concentration of axonal RNAs, localization in specific cortical domains, and the need to isolate axons. We demonstrate the importance of axon micro-dissection to obtain a whole mount perspective of mRNA distribution in the axonal territory. We describe a protocol to perform fluorescent ISH in isolated axons and guidelines for the preservation of structural and molecular integrity of cortical RNA-containing domains (e.g., Periaxoplasmic Ribosomal Plaques, or PARPs) in isolated axoplasm.
authors: Department of Proteins and Nucleic Acids, Instituto de Investigaciones Biológicas Clemente Estable (IIBCE), Av. Italia 3318, CP 11600, Montevideo, Uruguay.
authors: José Roberto
authors: JR
authors: Sotelo-Silveira
authors: Aldo
authors: A
authors: Calliari
authors: Alejandra
authors: A
authors: Kun
authors: Victoria
authors: V
authors: Elizondo
authors: Lucía
authors: L
authors: Canclini
authors: José Roberto
authors: JR
authors: Sotelo
mesh_terms: D000818:Animals; D001369:Axons / Q000378:metabolism*; D002469:Cell Separation; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D051379:Mice; D009186:Myelin Sheath / Q000502:physiology; D015345:Oligonucleotide Probes / Q000235:genetics; D034443:RNA Transport; D012333:RNA, Messenger / Q000032:analysis* / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D013126:Spinal Nerve Roots / Q000166:cytology / Q000378:metabolism; D016707:Tissue Fixation
chemical_list: D015345:Oligonucleotide Probes; D012333:RNA, Messenger