Genetius

Using CRISPR/Cas9 genome editing in Arabidopsis thaliana, the authors dissected a 2.3‑kb downstream region of the FLOWERING LOCUS T (FT) gene and identified the Block E enhancer as essential for proper FT expression and flowering. Fine‑scale deletions pinpointed a 63‑bp core module containing CCAAT‑ and G‑boxes, while a cryptic CCAAT‑box becomes active when repositioned, highlighting the importance of motif context and local chromatin accessibility for enhancer function.

The authors created a CRISPR/Cas9‑induced restoration bioluminescence reporter system (CiRBS) that reactivates an inactive luciferase mutant in the genome of transgenic Arabidopsis, enabling long‑term single‑cell bioluminescence monitoring. Rapid indel‑mediated recombination restores luciferase activity within 24 h, with ~7 % of transfected cells regaining bioluminescence, allowing reliable analysis of cellular gene‑expression heterogeneity from a single genomic locus.

The study applied a CRISPR/Cas9 multiplex guide RNA strategy to delete entire open reading frames of four reproductive genes in Arabidopsis thaliana, achieving homozygous deletions already in the T1 generation with rates of 8.3–30%. Deletion efficiencies correlated with DeepSpCas9 prediction scores, and phenotypic analyses revealed unexpected effects of residual gene fragments on fertilization and seed development.