Isolation of tissues and preservation of RNA from intact, germinated barley grain

The Plant Journal : for Cell and Molecular Biology
Natalie S BettsG B Fincher

Abstract

Isolated barley (Hordeum vulgare L.) aleurone layers have been widely used as a model system for studying gene expression and hormonal regulation in germinating cereal grains. A serious technological limitation of this approach has been the inability to confidently extrapolate conclusions obtained from isolated tissues back to the whole grain, where the co-location of several living and non-living tissues results in complex tissue-tissue interactions and regulatory pathways coordinated across the multiple tissues. Here we have developed methods for isolating fragments of aleurone, starchy endosperm, embryo, scutellum, pericarp-testa, husk and crushed cell layers from germinated grain. An important step in the procedure involves the rapid fixation of the intact grain to freeze the transcriptional activity of individual tissues while dissection is effected for subsequent transcriptomic analyses. The developmental profiles of 19 611 gene transcripts were precisely defined in the purified tissues and in whole grain during the first 24 h of germination by RNA sequencing. Spatial and temporal patterns of transcription were validated against well-defined data on enzyme activities in both whole grain and isolated tissues. Transcript pr...Continue Reading

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Citations

Oct 20, 2019·The Plant Journal : for Cell and Molecular Biology·Lim Chee LiewMathew G Lewsey
Mar 11, 2020·Foods·Daniel RicoAna B Martín-Diana
Dec 17, 2017·Journal of Integrative Plant Biology·Wai Li LimRachel A Burton
May 1, 2019·International Journal of Molecular Sciences·Mercedes Diaz-MendozaManuel Martinez
Dec 11, 2019·Journal of Experimental Botany·Natalie S BettsGeoffrey B Fincher
Feb 2, 2021·Analytical Biochemistry·Yuri F DryginTatiana V Gasanova
May 27, 2021·Plant Science : an International Journal of Experimental Plant Biology·Natalie S BettsVincent Bulone

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