Targeted gene inactivation in petunia by PCR-based selection of transposon insertion mutants

Proc Natl Acad Sci U S A. 1995 Aug 29;92(18):8149-53. doi: 10.1073/pnas.92.18.8149.

Abstract

Establishment of loss-of-function phenotypes is often a key step in determining the biological function of a gene. We describe a procedure to obtain mutant petunia plants in which a specific gene with known sequence is inactivated by the transposable element dTph1. Leaves are collected from batches of 1000 plants with highly active dTph1 elements, pooled according to a three-dimensional matrix, and screened by PCR using a transposon- and a gene-specific primer. In this way individual plants with a dTph1 insertion can be identified by analysis of about 30 PCRs. We found insertion alleles for various genes at a frequency of about 1 in 1000 plants. The plant population can be preserved by selfing all the plants, so that it can be screened for insertions in many genes over a prolonged period.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alleles
  • Base Sequence
  • DNA Primers
  • DNA Transposable Elements*
  • Gene Expression Regulation, Plant / genetics
  • Genes, Plant
  • Molecular Sequence Data
  • Mutagenesis, Insertional*
  • Plants / genetics*
  • Polymerase Chain Reaction

Substances

  • DNA Primers
  • DNA Transposable Elements