A method for obtaining high quality RNA from paraffin sections of plant tissues by laser microdissection

Hirokazu Takahashi, Hisae Kamakura, Yutaka Sato, Katsuhiro Shiono, Tomomi Abiko, Nobuhiro Tsutsumi, Yoshiaki Nagamura, Naoko K. Nishizawa, Mikio Nakazono

Research output: Contribution to journalArticle

70 Citations (Scopus)

Abstract

Laser microdissection (LM) combined with microarray analysis or next-generation sequencing of cDNA is a powerful tool for understanding molecular events in individual cell types of plants as well as animals. Obtaining high quality RNA is essential for this approach. For plant tissues, paraffin-embedded sections better preserve cell structure than do frozen sections. However, the conventional method for preparing paraffin sections is a lengthy process involving embedding the tissue and floating and drying the sections, during which time RNA degradation occurs. Here, we describe a method for preparing serial sections that greatly reduces RNA degradation: we reduced (1) the embedding time from 4-6 days to about 5 h by using a recently developed microwave method; (2) the time of floating sections from ~10 min to less than 5 min, (3) the drying time from ~12 to 1 h; and (4) the drying temperature from 42 to 4°C. With this method, we were able to isolate higher integrity RNA from many kinds of plant tissues than is typically obtained by the conventional paraffin preparation method. The improvement in RNA quality and yield removes a major obstacle to the widespread use of LM with high-throughput technologies for plants.

Original languageEnglish
Pages (from-to)807-813
Number of pages7
JournalJournal of Plant Research
Volume123
Issue number6
DOIs
Publication statusPublished - Jun 1 2010
Externally publishedYes

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alkanes
lasers
plant tissues
RNA
drying
methodology
degradation
cell structures
drying temperature
preserves
animals
cells

All Science Journal Classification (ASJC) codes

  • Plant Science

Cite this

A method for obtaining high quality RNA from paraffin sections of plant tissues by laser microdissection. / Takahashi, Hirokazu; Kamakura, Hisae; Sato, Yutaka; Shiono, Katsuhiro; Abiko, Tomomi; Tsutsumi, Nobuhiro; Nagamura, Yoshiaki; Nishizawa, Naoko K.; Nakazono, Mikio.

In: Journal of Plant Research, Vol. 123, No. 6, 01.06.2010, p. 807-813.

Research output: Contribution to journalArticle

Takahashi, H, Kamakura, H, Sato, Y, Shiono, K, Abiko, T, Tsutsumi, N, Nagamura, Y, Nishizawa, NK & Nakazono, M 2010, 'A method for obtaining high quality RNA from paraffin sections of plant tissues by laser microdissection', Journal of Plant Research, vol. 123, no. 6, pp. 807-813. https://doi.org/10.1007/s10265-010-0319-4
Takahashi, Hirokazu ; Kamakura, Hisae ; Sato, Yutaka ; Shiono, Katsuhiro ; Abiko, Tomomi ; Tsutsumi, Nobuhiro ; Nagamura, Yoshiaki ; Nishizawa, Naoko K. ; Nakazono, Mikio. / A method for obtaining high quality RNA from paraffin sections of plant tissues by laser microdissection. In: Journal of Plant Research. 2010 ; Vol. 123, No. 6. pp. 807-813.
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