Genetic engineering of wheat for polyamine modification

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A facile method for screening for phosphinothricin (PPT)-resistant transgenic wheats

A bioassay was developed for identifying transgenic wheat plants based on De Block et al.'s [8] ammonium-multiwell assay which allows qualitative and quantitative evaluation of the expression of the enzyme phosphinothricin acetyl transferase (PAT). Important parameters in the assay are the use of young leaf tissues, short incubation period (6 h) and a high light intensity during incubation (250 μmol s−1m−2). The assay is quick and results are obtained within a day. Ammonium measurements based on a colourimetric (modified Berthelot) reaction are conducted using sodium salicylate to avoid the use of phenol. Results of the assay show high correlation with Basta leaf painting tests and polymerase chain reaction (PCR) results. Thus, the assay may be used as a facile screen for bar-expressing transgenic cereals

Analysis of particle bombardment parameters to optimise DNA delivery into wheat tissues

The objective of this study was to identify the major parameters controlling DNA delivery by particle bombardment to wheat (Triticum aestivum L.) scutellum and inflorescence tissue. The main factors studied were the DNA/gold precipitation process, bombardment parameters and tissue culture variables. Efficiency of DNA (uidA gene) delivery was assessed by scoring transient GUS expression in bombarded tissues. Of the parameters analysed, amount of plasmid DNA, spermidine concentration, presence of Ca++ ions, calcium chloride concentration, amount of gold particles, gold particle size, acceleration pressure, chamber vacuum pressure, bombardment distance, osmotic conditioning of tissues and type of auxin had a clear influence on transient gene expression. A bombardment procedure suitable for elite wheat varieties was developed which allowed high-efficiency DNA delivery combined with reduced damage to target tissues