Gene trapping with firefly luciferase in Arabidopsis. Tagging of stress-responsive genes

Plant Physiol. 2004 Jan;134(1):18-27. doi: 10.1104/pp.103.027151.

Abstract

To monitor the expression of T-DNA-tagged plant genes in vivo, a collection of 20,261 transgenic lines of Arabidopsis (Columbia-0) were generated with the promoter trap vector pTluc, which carries a promoterless firefly luc (luciferase) reporter gene linked to the right T-DNA border. By detection of bioluminescence in 3-week-old seedlings, 753 lines were identified showing constitutive, organ-specific, and stress-responsive luciferase expression patterns. To facilitate the identification of well-defined luciferase expression patterns, a pooled seed stock was established. Several lines showed sugar, salt, and abscisic acid (ABA)-inducible luciferase activity. Segregation analysis of 215 promoter trap lines indicated that about 50% of plants contained single insertions, whereas 40% carried two and 10% carried three or more T-DNA tags. Sequencing the T-DNA insert junctions isolated from 17 luciferase-expressing lines identified T-DNA tags in 5'- and 3'-transcribed domains and translational gene fusions generated by T-DNA insertions in exons and introns of Arabidopsis genes. Tissue specific expression of eight wild-type Arabidopsis genes was confirmed to be similar to the luminescence patterns observed in the corresponding luciferase-tagged lines. Here, we describe the characterization of a transcriptional luc reporter gene fusion with the WBC-type ABC transporter gene At1g17840. Expression of wild-type and luciferase-tagged At1g17840 alleles revealed similar induction by salt, glucose, and ABA treatments and gibberellin-mediated down-regulation of ABA-induced expression. These results illustrate that luciferase gene traps are well suited for monitoring the expression of stress-responsive Arabidopsis genes in vivo.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Arabidopsis / genetics*
  • Artificial Gene Fusion
  • Base Sequence
  • Coleoptera / enzymology
  • Coleoptera / genetics
  • DNA, Bacterial / genetics
  • Gene Expression
  • Genes, Plant
  • Genes, Reporter
  • Genetic Vectors
  • Luciferases / genetics*
  • Molecular Sequence Data
  • Plants, Genetically Modified
  • Promoter Regions, Genetic
  • Recombinant Proteins / genetics

Substances

  • DNA, Bacterial
  • Recombinant Proteins
  • T-DNA
  • Luciferases