Cloning And Expression Of Two ICE1 Homologous Gene From Vitis Amurensis

The low temperature is limiting important environmental factor of geographical dist of plants and growth, moreover the low temperature freeze injury is the global agriculture forestry universal existence natural disaster, has created the massive loss for the world agriculture forestry. Along with the molecular biology technology's development, the winter-killing research already penetrated into the molecular level, thus promotion cold resistant of plant genetic engineering development.This experiment take the amur grape as material,Two ICE1 genes were isolated from Vitis amurensis Rupr.by bioformatic analysis and RT-PCR. named as VaICE1a and the VaICE1b gene. The result is as follows:VaICE1a gene cDNA span is 1617bp, according to the nucleotide analysis that it was composed 538 amino acids, named VaICE1a. The cDNA sequence and the code protein sequence showed that it includes a rich s area, the nucleus localization signaling zone (NLS), to transfer the membrane area. Homology analysis showed that the deduced VaICE1 protein was highly homologous to the Vitis vinifera (XP-002275711),Malus,Populus trichocarpa,Vitis vinifera (XP-002284528),Arabidopsis thaliana,Triticum aestivum, the identity are 99%,61%,62%,64%,60%,76%, respectively. Sequence in C terminal is high conserved, and has bHLH typical structure. Phylogenetic analysis also indicated that VaICE1b belongs to the ICE1 gene family. Tertiary Structure analysis that is mixed protein. The above result indicated that VaICE1a is possibly a ICE1 gene family's young blood.Semi-quantitative PCR results showed that VaICE1a gene indifferent tissues of Vitis amurensis Rupr. was expressed degrees. under abiotic Stress, VaICE1a gene could induce expresses, The expression Level was not obvious difference,compare with control groupes.VaICE1b gene cDNA span is 1653bp, according to the nucleotide analysis that it was composed 550amino acids, named VaICE1b. The cDNA sequence and the code protein sequence showed that it includes a rich s area, the nucleus localization signaling zone (NLS), to transfer the membrane area. Homology analysis showed that the deduced VaICE1 protein was highly homologous to the Vitis vinifera (XP-002275711). Malus,Populus trichocarpa,Vitis vinifera (XP-002284528),Arabidopsis thaliana,Triticum aestivum, the identity are 64%,68%,68%,99%,52%,73%, respectively. Sequence in C terminal is high conserved, and it has bHLH typical structure. Phylogenetic analysis also indicated that VaICE1b belongs to the ICE1 gene family. Tertiary Structure analysis that it is mixed protein. The above result indicated that VaICE1a is possibly a ICE1 gene family's new member. Semi-quantitative PCR results showed that VaICE1b gene indifferent tissues of Vitis amurensis Rupr. was expressed degrees. The expression is relative highly compare with control groupes.We constructed plant overexpression vector of VaICE1a of VaICE1b.