Prior to the mapping and deep sequencing a mapping population of plants was prepared by first crossing the mutant lineages of Columbia ecotype with wild type plants of the Landsberg ecotype. This generation was allowed to self fertilize to give rise to the mapping population. Homozygous mutants from the mapping population was then analysed using single sequence length polymorphism (SSLP) markers being able to separate between Columbia and Landsberg background. The markers were used to retrieve information about recombinations in the area of interest, where no recombinations were associated with homozygosity for Col, one recombination with a heterozygous Col/Ler genotype and two recombinations with homozygosity for Ler. As the mutations were introduced in a Columbia background the area surrounding the mutation of the homozygous mutants would also be of the Columbia ecotype. When two markers surrounding the mutation had been found the area between them was investigated using a third marker narrowing down the area of interest by three-point mapping. Mutations were located on lower arm of chromosome IV and upper arm of chromosome I respectively.

At this point locating and identifying genes carrying mutations proceeded with deep sequencing, even if traditional mapping could have been used as well. A total of four mutants were sent to Max Planck Institute, Tübingen, Germany, for deep sequencing. Two were the above mentioned ectopic expressors and the other two were low/no-signal mutants. The deep sequencing confirmed the mapping results for the ectopic expressors and also found accumulation of Columbia ecotype on upper and lower arms of chromosome V in the two low/no-signal mutants. Single-nucleotide polymorphisms (SNPs) were identified in those regions and genes carrying SNPs causing amino acid exchange were chosen for further examinations. Three really interesting genes were found and they encoded:

• A protein involved in synthesis of polyamines. Polyamines have been shown to prevent premature cell death in immature vessels by suppressing genes involved in PCD.

• A protein involved in the biosynthesis of the plant hormone abscisic acid. Abscisic acid regulates the metabolism of polyamines in response to abiotic stress.

• A protein important in lignin biosynthesis. Lignin is an important constituent of the vessel walls as it makes the vessels rigid.