title: A method for extracting high-quality RNA from diverse plants for next-generation sequencing and gene expression analyses.
abstract: OBJECTIVE
  To study gene expression in plants, high-quality RNA must be extracted in quantities sufficient for subsequent cDNA library construction. Field-based collections are often limited in quantity and quality of tissue and are typically preserved in RNAlater. Obtaining sufficient and high-quality yield from variously preserved samples is essential to studies of comparative biology. We present a protocol for the extraction of high-quality RNA from even the most recalcitrant plant tissues. •
  
  
  RESULTS
  Tissues from mosses, cycads, and angiosperm floral organs and leaves were preserved in RNAlater or frozen fresh at -80°C. Extractions were performed and quality was measured for yield and purity. •
  
  
  CONCLUSIONS
  This protocol results in the extraction of high-quality RNA from a variety of plant tissues representing vascular and nonvascular plants. RNA was used for cDNA synthesis to generate libraries for next-generation sequencing and for expression studies using quantitative PCR (qPCR) and semiquantitative reverse transcription PCR (RT-PCR).
authors.affiliation: Department of Plant and Microbial Biology, Department of Integrative Biology, and the University and Jepson Herbaria, University of California, Berkeley, California 94720 USA ; Origine, Structure et Evolution de la Diversité (UMR 7205 CNRS), Muséum National d'Histoire Naturelle, CP39, 16 rue Buffon, 75231 Paris Cedex 05, France.
authors.forename: Roxana
authors.initials: R
authors.lastname: Yockteng
authors.affiliation: Department of Plant and Microbial Biology, Department of Integrative Biology, and the University and Jepson Herbaria, University of California, Berkeley, California 94720 USA.
authors.forename: Ana M R
authors.initials: AM
authors.lastname: Almeida
authors.affiliation: Department of Plant and Microbial Biology, Department of Integrative Biology, and the University and Jepson Herbaria, University of California, Berkeley, California 94720 USA.
authors.forename: Stephen
authors.initials: S
authors.lastname: Yee
authors.affiliation: Laboratório de Taxonomia Vegetal e Biologia Reprodutiva, Departamento de Botânica, Universidade Federal do Rio de Janeiro, 21941-902 Rio de Janeiro, Rio de Janeiro, Brazil.
authors.forename: Thiago
authors.initials: T
authors.lastname: Andre
authors.affiliation: Department of Plant and Microbial Biology, Department of Integrative Biology, and the University and Jepson Herbaria, University of California, Berkeley, California 94720 USA.
authors.forename: Colin
authors.initials: C
authors.lastname: Hill
authors.affiliation: Department of Plant and Microbial Biology, Department of Integrative Biology, and the University and Jepson Herbaria, University of California, Berkeley, California 94720 USA.
authors.forename: Chelsea D
authors.initials: CD
authors.lastname: Specht
keywords: Illumina; RNA extraction; RNA-Seq; cDNA library; gene expression; transcriptome