Construction of a cDNA library of the Chinese wild Vitis amurensis under cold stress and analysis of potential hardiness-related expressed sequence tags.

A cDNA library of Chinese wild Vitis amurensis, which is the most cold-resistant species in the genus Vitis, was constructed using young leaves of seedlings of the clone Heilongjiang potted and subjected to cold stress. The leaves were harvested at various times after cold stress for total RNA extraction, which was used to generate expressed sequence tags (ESTs). The titer of the original library was 2.67 x 10(6) pfu/mL and the corresponding combination frequency was 98.5%. The test values of the amplified library were 1.53 x 10(9) pfu/mL and 98.2%, respectively. After randomly choosing, cloning and sequencing, 227 ESTs with high quality were obtained and submitted to GenBank database. Using BLASTX, 79.7% of the ESTs shared homology with known functional DNA sequences and 20.3% shared significant homology with unknown proteins. Some potential hardiness-related ESTs were obtained, which were involved in signal transduction, stress inducement, defense reactions, transcription factors, etc. Some functional genes identified from the cDNA library have potential for plant defense. These sequences will be subjected to further researches on hardiness genes and their molecular mechanisms.