Testing for frost tolerance in wheat (Triticum aestivum L.) transformed with a gene for antifreeze protein

Abstract

Spring frosts in wheat growing areas of Australia can result in significant damage during the heading stage, resulting in losses millions of dollars each year due to decreased yields. To address this issue, wheat has been co-transformed via micro-projectile bombardment with an antifreeze protein (AFP) gene targeted to the apoplast or endoplasmic reticulum. T0 transgenic plants expressed AFP and showed the potential for frost tolerance in in vitro tests on plant extracts (Khanna and Daggard 2005). Additional transgenic wheat plants carrying the AFP gene targeted to both apoplast and ER have subsequently been produced. DNA extracted from plants surviving selection on phosphinothricin revealed 63.8% were PCR positive for presence of the AFP gene, indicating a wheat transformation frequency of 3.97%. Co-transformation of AFP and bar genes occurred in 53% of samples tested. PCR and gene expression analysis (via RT-PCR) have been carried out on plants that were subsequently tested in a controlled frost environment at the Australian Genome Research Facility (AGRF Adelaide). Analysis of frost induced sterility (FIS) showed that frosted control plants were significantly higher in FIS compared to unfrosted controls, and that FIS of plants transformed with one of the constructs was not significantly different to that of the unfrosted controls. This result suggests that the introduced AFP may protect the transgenic wheat plants against frost.