Cell Wall Epitopes and Endoploidy as Reporters of Embryogenic Potential in Brachypodium Distachyon Callus Culture

Alexander Betekhtin, Magdalena Rojek, Katarzyna Nowak, Artur Pinski, Anna Milewska-Hendel, Ewa Kurczynska, John Doonan, Robert Hasterok

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Abstract

Effective regeneration of callus tissue into embryos and then into whole plants is essential for plant biotechnology. The embryonic potential is often low and can further decrease with time in culture, which limits the utilisation of calli for transformation procedures and in vitro propagation. In this study, we show that the loss of embryogenic potential in callus cultures of Brachypodium distachyon is progressive over time. Flow cytometry analyses indicated endoploidy levels increased in 60- and 90-day-old calli with effective loss of the 2C DNA content peak in the latter. Analysis of indolic compounds content revealed a decrease in 60- and 90-day-old calli compared to either freshly isolated explants or 30-day-old calli. Immunohistochemical analysis revealed a decrease in arabinogalactan proteins (AGP) signal with the time of culture, but extensin (EXT) epitopes either increased (JIM12 epitopes) or decreased (JIM11 epitopes). The transcript accumulation levels of AGPs and EXTs confirmed these results, with most of AGP and EXT transcripts gradually decreasing. Some chimeric EXT transcripts significantly increased on the 30th day of culture, perhaps because of an increased embryogenic potential. Selected somatic embryogenesis-related genes and cyclins demonstrated a gradual decrease of transcript accumulation for YUCCA (YUC), AINTEGUMENTA-LIKE (AIL), BABY BOOM (BBM), and CLAVATA (CLV3) genes, as well as for most of the cyclins, starting from the 30th day of culture. Notably, WUSCHEL (WUS) transcript was detectable only on the 30th and 60th day and was not detectable in the zygotic embryos and in 90-day-old calli
Original languageEnglish
Article number3811
JournalInternational Journal of Molecular Sciences
Volume19
Issue number12
DOIs
Publication statusPublished - 29 Nov 2018

Keywords

  • arabinogalactan proteins
  • Brachypodium distachyon
  • cell wall
  • cyclins
  • extensins
  • pectins
  • ploidy instability
  • somatic embryogenesis
  • somaclonal variation
  • transcript accumulation
  • Somatic embryogenesis
  • Extensins
  • Somaclonal variation
  • Cell wall
  • Cyclins
  • Transcript accumulation
  • Arabinogalactan proteins
  • Ploidy instability
  • Pectins
  • Mucoproteins/metabolism
  • Epitopes/immunology
  • Embryonic Development/physiology
  • Plant Somatic Embryogenesis Techniques
  • Brachypodium/cytology
  • Flow Cytometry
  • Glycoproteins/metabolism
  • Plant Proteins/metabolism
  • Pectins/metabolism
  • Cell Wall/metabolism
  • Cyclins/metabolism

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