Cold Tolerance of Transgenic Tobacco Carrying Gene Encoding Insect Antifreeze Protein

Abstract

The MPAFP149 gene from Xinjiang desert insect Microdera punctipennis dzhungarica was constructed to the plant expression vector pCAMBIA 1302 by sub-cloning. The recombined vector pCAMBIAI3O2-MPAFPI49 was transformed into Agrobacterium tumefaciens EHA 105. Transgenic tobacco was obtained via leaf-disc method by Agrobacterium-mediated transformation. PCR and PCR-Southern analysis showed that the MPAFPI49 gene was successfully integrated into the tobacco genome. The result of RT-PCR also verified that the MPAFPI49 gene was transcribed at the mRNA level. Transgenic tobacco showed greater cold tolerance than wild tobacco. At room temperature, the transgenic tobacco recovered from cold stress and regenerated. The results confirmed that transgenic tobacco carrying the MPAFP149 gene was more tolerant of cold than wild tobacco.