Rapid analysis of circadian phenotypes in Arabidopsis protoplasts transfected with a luminescent clock reporter

Louise L. Hansen, Gerben Van Ooijen

Research output: Contribution to journalArticlepeer-review

Abstract / Description of output

The plant circadian clock allows the anticipation of daily changes to the environment. This anticipation aids the responses to temporally predictable biotic and abiotic stress. Conversely, disruption of circadian timekeeping severely compromises plant health and reduces agricultural crop yields. It is therefore imperative that we understand the intricate regulation of circadian rhythms in plants, including the factors that affect motion of the transcriptional clockwork itself.

Testing circadian defects in the model plant Arabidopsis thaliana (Arabidopsis) traditionally involves crossing specific mutant lines to a line rhythmically expressing firefly luciferase from a circadian clock gene promoter. This approach is laborious, time-consuming, and could be fruitless if a mutant has no circadian phenotype. The methodology presented here allows a rapid initial assessment of circadian phenotypes.

Protoplasts derived from mutant and wild-type Arabidopsis are isolated, transfected with a rhythmically expressed luminescent reporter, and imaged under constant light conditions for 5 days. Luminescent traces will directly reveal whether the free-running period of mutant plants is different from wild-type plants. The advantage of the method is that any Arabidopsis line can efficiently be screened, without the need for generating a stably transgenic luminescent clock marker line in that mutant background.
Original languageEnglish
Article numbere54586
Number of pages7
JournalJournal of Visualized Experiments (JoVE)
Early online date17 Sept 2016
Publication statusPublished - 17 Sept 2016

Keywords / Materials (for Non-textual outputs)

  • circadian clocks
  • chronobiology
  • Arabidopsis thaliana
  • protoplasts
  • Luminescent imaging
  • transfection
  • clock genes
  • phenotyping
  • Plant Biology


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