Establishment of Rhizobium-mediated transformation of Coptis japonica and molecular analyses of transgenic plants

Nobukazu Shitan, Fumiyuki Kiuchi, Fumihiko Sato, Kazufumi Yazaki, Kayo Yoshimatsu

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

Coptis japonica is a perennial medicinal plant grown in Asia, and its rhizome is used as crude drug, in which berberine is highly accumulated as the main alkaloid. In this study, a transformation method for C. japonica using Rhizobium has been established. Young petioles were infected with Rhizobium radiobactor Gv3101 (pMP90) harboring a plasmid with β-glucuronidase (gus) driven by cauliflower mosaic virus 35S promoter and hygromycin phosphotransferase (hpt) gene as the selection marker. GUS assay showed that 46% of the calli grown on hygromycin plates clearly expressed the gus gene. As an application of the transformation method, an endogenous cDNA encoding an ABC protein, CjMDR1, which is involved in berberine transport, was introduced in sense orientation. Cjmdr1 transgenic plant (Sense-2) regenerated from hygromycin resistant callus line had a single copy of integrated hpt gene in its genomic DNA. Northern analyses of the Sense-2 plant showed that Cjmdr1 mRNA levels were lower in all organs examined than those of wild-type plant, which suggested that co-suppression of Cjmdr1 expression occurred. Indeed, the berberine content in Sense-2 was also suppressed. This is the first report of stable transformation of Coptis spp. and also of the alteration of a secondary metabolite by transport engineering.

Original languageEnglish
Pages (from-to)113-118
Number of pages6
JournalPlant Biotechnology
Volume22
Issue number2
Publication statusPublished - 2005 Jun
Externally publishedYes

Fingerprint

hygromycin-B kinase
Coptis japonica
Coptis
Berberine
berberine
Rhizobium
Genetically Modified Plants
transgenic plants
Glucuronidase
Bony Callus
callus
phosphotransferases (kinases)
Caulimovirus
Genes
Rhizome
genes
Cauliflower mosaic virus
Medicinal Plants
petioles
RNA Interference

Keywords

  • Berberine
  • Coptis japonica
  • GUS expression
  • Rhizobium radiobacter

ASJC Scopus subject areas

  • Plant Science

Cite this

Establishment of Rhizobium-mediated transformation of Coptis japonica and molecular analyses of transgenic plants. / Shitan, Nobukazu; Kiuchi, Fumiyuki; Sato, Fumihiko; Yazaki, Kazufumi; Yoshimatsu, Kayo.

In: Plant Biotechnology, Vol. 22, No. 2, 06.2005, p. 113-118.

Research output: Contribution to journalArticle

Shitan, Nobukazu ; Kiuchi, Fumiyuki ; Sato, Fumihiko ; Yazaki, Kazufumi ; Yoshimatsu, Kayo. / Establishment of Rhizobium-mediated transformation of Coptis japonica and molecular analyses of transgenic plants. In: Plant Biotechnology. 2005 ; Vol. 22, No. 2. pp. 113-118.
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