Antifreeze proteins (AFPs) enable organisms to survive under cold conditions, and have great potential in improv- ing cold tolerance of cold-sensitive plants. In order to determine whether expression of the carrot 36 kD antifreeze protein gene con- fers improved cold-resistant properties to plant tissues, we tried to obtain transgenic tobacco plants which expressed the antifreeze protein. Cold, salt, and drought induced promoter Prd29A was cloned using PCR from Arabidopsis. Two plant expression vectors based on pBI121 were constructed with CaMV35S:AFP and Prd29A:AFP. Tobacco plantlets were transformed by Agrobacte- rium-medicated transformation. PCR and Southern blotting demonstrated that the carrot 36 kD afp gene was successfully integrated into the genomes of transformed plantlets. The expression of the afp gene in transgenic plants led to improved tolerance to cold stress. However, the use of the strong constitutive 35S cauliflower mosaic virus (CaMV) promoter to drive expression of afp also resulted in growth retardation under normal growing conditions. In contrast, the expression of afp driven by the stress-inducible Prd29A pro- moter from Arabidopsis gave rise to minimal effects on plant growth while providing an increased tolerance to cold stress condition (2°C). The results demonstrated the prospect of using Prd29A-AFP transgenic plants in cold-stressed conditions that will in turn benefit agriculture.
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