Differentially Expressed cDNAs in Cold-Stressed SSH Library of Ceratoides lanata

As a high stress resistance psammophyte with the characteristic of sand fixation and winterfat, Ceratoides lanata had been successfully introduced into many countries in recent years. The leaves and twigs of Ceratoides lanata in room temperature were used as the Driver and those treated by cold stress were used as Tester. Tester and Driver ds cDNA were prepared from their high quality mRNA which were purified from their total RNA. Tester and Driver cDNA were separately digested to obtain shorter, blunt-ended molecules by RasI. Then two Tester populations are created with different adaptors, while Driver cDNA has no adaptors. Differentially expressed genes were equalized and enriched by two round subtractive hybridizations using excess Driver population as compared with Tester population. The differentially expressed cDNAs were exponentially amplified by first round suppression PCR using the diluted hybridization product as template, and then the secondary PCR was performed using the first PCR product as template by nested primers to finally enrich the differentially expressed cDNAs, which consists of the SSH library of Ceratoides lanata. These cDNAs were inserted into vectors and 362 cDNA clones were obtained. The sequencing results showed that some cDNAs of cold-stressed Ceratoides lanata SSH library have relatively high homology with known stress resistance-related genes or proteins, and other cDNAs are new genes which are firstly reported here. Our result lays a foundation for the cDNA cloning of valuable genes including antifreeze genes, heat-resistant genes, drought-tolerant genes, alkali-salt-tolerant genes from Ceratoides lanata, and their transgenic application.