Genetic and phenotypic analyses of a papaver somniferum T-DNA insertional mutant with altered alkaloid composition

Noriaki Kawano, Fumiyuki Kiuchi, Nobuo Kawahara, Kayo Yoshimatsu

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

The in vitro shoot culture of a T-DNA insertional mutant of Papaver somniferum L. established by the infection of Agrobacterium rhizogenes MAFF03-01724 accumulated thebaine instead of morphine as a major opium alkaloid. To develop a non-narcotic opium poppy and to gain insight into its genetic background, we have transplanted this mutant to soil, and analyzed its alkaloid content along with the manner of inheritance of T-DNA insertion loci among its selfed progenies. In the transplanted T0 primary mutant, the opium (latex) was found to be rich in thebaine (16.3% of dried opium) by HPLC analysis. The analyses on T-DNA insertion loci by inverse PCR, adaptor-ligation PCR, and quantitative real-time PCR revealed that as many as 18 copies of T-DNAs were integrated into a poppy genome in a highly complicated manner. The number of copies of T-DNAs was decreased to seven in the selected T3 progenies, in which the average thebaine content was 2.4-fold that of the wild type plant. This may indicate that the high thebaine phenotype was increasingly stabilized as the number of T-DNA copies was decreased. In addition, by reverse transcription PCR analysis on selected morphine biosynthetic genes, the expression of codeine 6-O-demethylase was clearly shown to be diminished in the T0 in vitro shoot culture, which can be considered as one of the key factors of altered alkaloid composition.

Original languageEnglish
Pages (from-to)133-154
Number of pages22
JournalPharmaceuticals
Volume5
Issue number2
DOIs
Publication statusPublished - 2012

Fingerprint

Papaver
Thebaine
Alkaloids
Opium
Polymerase Chain Reaction
Morphine
O Demethylating Oxidoreductases
Codeine
Agrobacterium
Latex
Reverse Transcription
Ligation
T-DNA
Real-Time Polymerase Chain Reaction
Soil
High Pressure Liquid Chromatography
Genome
Phenotype
Gene Expression
Infection

Keywords

  • Agrobacterium rhizogenes
  • Opium alkaloid
  • Papaver somniferum L.
  • T-DNA insertional mutant

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmaceutical Science

Cite this

Genetic and phenotypic analyses of a papaver somniferum T-DNA insertional mutant with altered alkaloid composition. / Kawano, Noriaki; Kiuchi, Fumiyuki; Kawahara, Nobuo; Yoshimatsu, Kayo.

In: Pharmaceuticals, Vol. 5, No. 2, 2012, p. 133-154.

Research output: Contribution to journalArticle

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