| Tamarix hispida, a woody halophyte, thrives in saline or saline-alkali soil, having a high ability in resisting saline and saline-alkali stress. The expression profiling in response to salinity-alkali stress in T.hispida was studied by large-scale expressed sequence tags (EST) analysis, and some full-length cDNA sequence of stress resistance gens were cloned from T.hispida and further analyzed.Three cDNA libraries were constructed from leaf tissue of T. hispida plants that were well-watered (control) or exposed to 0.4 mol/L NaHCO3 for 24 and 52 h (the three libraries were signed as 0,24 and 52 h library, respectively). The primary titer of the three libraries were 7.5x105 pfu, 2.6×106 pfu and 4.0×106 pfu,, with the average size of inserts of 1.1, 1.2 and 1.2 kb, respecitvly. The average rate of recombination in three libraries were all about 98%.A total of 9447 high quality expressed sequence tags (ESTs) were obtained from the three libraries. They were submitted to the GenBank databases with accession numbers from EH048198 to EH057644. The 9447 ESTs were assembled into 986 contigs and 2959 singlets. The numbers of Unigenes from the 0, 24 and 52 h library were 1752, 1558 and 1675, respectively.Among them, 7365 ESTs (77.96%) showed significant similarities (E-value＜10-4) to gene sequences in the non-redundant (Nr) GenBank database, and the other 2082 (22.04%) had little, or no, similarity. The database-matched ESTs were further grouped into 12 funtion categories, i.e., photosynthesis (20.31%), function unknown (13.97%), cell rescue, defense (13.96%), energy (7.59%), protein synthesis (7.58%), transport facilitation (7.28%), transcription (6.63%), cell structure (6.07%), metabolism-related category (5.70%), protein destination (5.02%), signal transduction (3.86%), and cell growth and division (2.04%).EST analysis was performed to compare gene expression in the three libraries, and the transcripts responsive to NaHCO3 were identified. We found the transcripts encoding osmolyte biosynthesis, hormone signaling transduction, scavenging reactive oxygen species, transcriptional regulators, protein synthesis and destination, ion homeostasis, ribosomal proteins, photosynthesis and metabolism were significantly regulated under NaHCO3 stress.To investigate the differences in gene regulation between T. hispida after exposure to NaC1 or NaHCO3, nine transcripts, differentially regulated by NaHCO3 stress, were selected for further expression analysis using real time reverse transcription-polymerase chain reaction (RT-PCR). Gene expression trends were similar after a 24-h exposure to either NaCl or NaHCO3, however, substantial variability was found after a 52-h exposure, indicating that short-term responses to either salt may not be significantly different. Bioinformation methods were conducted in cloning the full-length cDNA sequence genes from three libraries, and the full-length genes were obtained. Among these genes, the vacuolar-H+ ATPase subunit c,, vacuolar-H+-ATPase subunit c"and two unique glycine-rich RNA-binding protein genes were selected for further expression analysis using real-time RT-PCR. The expressions of ThVHA el and Th VHA c"l were significantly increased while the expressions of ThGRPl and ThGRP2 were continually decreased after exposure to NaC1. However, the Th VHA el and Th VHA c"l were both down-regulated after NaHCO3 treatment for 24 h and up- regulated for 52 h. The ThGRP1 and ThGRP2 genes were both up-regulated at 24 h and down- regulated at 52 h of NaHCO3 treatment.These sstudies showed that the response to saline-alkali stress in T. hispida is a complex one, involving multiple physiological and metabolic pathways. The results will enrich our knowledge of the stress tolerance of woody plants on a molecular level and provide new insight into saline-alkali tolerance in plants. In addition, the cloning and identification of saline-alkali tolerance genes and determination of their expression patterns under saline-alkali stress, may offer some attractive candidate genes and valuable information for improving saline-alkali tolerance of other woody plant species through genetic engineering.
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