title: High quality RNA extraction of the mammalian cochlea for qRT-PCR and transcriptome analyses.
abstract: Molecular investigations of the hearing organ, the cochlea, have been hampered due to the difficulty of isolating pure RNA and in quantities sufficient enough for quantitative real-time RT-PCR or microarray analysis. The complex architecture of the cochlea, the presence of liquids, bone and cartilage tissue, are a major hurdle in obtaining contamination-free RNA to a level that does not affect downstream applications. Here, we present a protocol to extract RNA from the mouse cochlea, with yields and quality suitable for real-time RT-PCR or Affymetrix labeling. In contrast to current methods, such as TRIZOL or column-based extraction, this protocol combines the two and, within 4 h, yields a 2 μg of total RNA from a single pair of adult mouse cochleae. This protocol allows the isolation of RNA molecules from the mammalian cochlea providing access to whole-transcript expression analyses.
authors.affiliation: Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, 17177, Sweden. Electronic address: kim.Vikhe.patil@stud.ki.se.
authors.forename: Kim
authors.initials: K
authors.lastname: Vikhe Patil
authors.affiliation: Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, 17177, Sweden. Electronic address: Barbara.Canlon@ki.se.
authors.forename: Barbara
authors.initials: B
authors.lastname: Canlon
authors.affiliation: Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, 17177, Sweden. Electronic address: christopher.cederroth@ki.se.
authors.forename: Christopher R
authors.initials: CR
authors.lastname: Cederroth
mesh_terms: D000818:Animals; D001842:Bone and Bones / Q000378:metabolism; D002356:Cartilage / Q000378:metabolism; D002725:Chloroform / Q000737:chemistry; D002940:Circadian Rhythm; D003051:Cochlea / Q000378:metabolism*; D004247:DNA / Q000737:chemistry; D004492:Edetic Acid / Q000737:chemistry; D020869:Gene Expression Profiling; D006146:Guanidines / Q000737:chemistry; D051379:Mice; D044085:Microfluidics; D020411:Oligonucleotide Array Sequence Analysis; D010636:Phenols / Q000737:chemistry; D012313:RNA / Q000737:chemistry*; D060888:Real-Time Polymerase Chain Reaction; D013053:Spectrophotometry
chemical_list: D006146:Guanidines; D010636:Phenols; C411644:trizol; D012313:RNA; D002725:Chloroform; D004247:DNA; D004492:Edetic Acid