Surrogate splicing for functional analysis of sesquiterpene synthase genes

Shuiqin Wu, Mark A. Schoenbeck, Bryan T. Greenhagen, Shunji Takahashi, Sungbeom Lee, Robert M. Coates, Joseph Chappell

Research output: Contribution to journalArticlepeer-review

27 Scopus citations


A method for the recovery of full-length cDNAs from predicted terpene synthase genes containing introns is described. The approach utilizes Agrobacterium-mediated transient expression coupled with a reverse transcription-polydeoxyribonucleotide chain reaction assay to facilitate expression cloning of processed transcripts. Subsequent expression of intronless cDNAs in a suitable prokaryotic host provides for direct functional testing of the encoded gene product. The method was optimized by examining the expression of an intron-containing β-glucuronidase gene agroinfiltrated into petunia (Petunia hybrida) leaves, and its utility was demonstrated by defining the function of two previously uncharacterized terpene synthases. A tobacco (Nicotiana tabacum) terpene synthase-like gene containing six predicted introns was characterized as having 5-epi-aristolochene synthase activity, while an Arabidopsis (Arabidopsis thaliana) gene previously annotated as a terpene synthase was shown to possess a novel sesquiterpene synthase activity for α-barbatene, thujopsene, and β-chamigrene biosynthesis.

Original languageEnglish
Pages (from-to)1322-1333
Number of pages12
JournalPlant Physiology
Issue number3
StatePublished - 2005

ASJC Scopus subject areas

  • Physiology
  • Genetics
  • Plant Science


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