Identification Of Genes Responsive To Na₂CO₃in The Roots Of Puccinellia Tenuiflora

Puccinellia tenuiflora, a halophyte with high capacity of salt tolerance, is anexcellent material for the separation and identification of salt tolerant genes and study ofmolecular mechanism of plant salt tolerance. In this study, cDNA Macroarraytechnology was used to screen differentially expressed clones among275clones from asuppression subtrative hybridization cDNA library constructed using Puccinelliatenuiflora roots grown on saline-alkali land. Bioinformatics analysis was carried out onthe screened differentially expressed genes. RT-PCR technology was performed tofurther investigate Na₂CO₃stress-responsive genes from the clones identified bymacroarray hybridization. The main results were as follows:1. Among the275clones from the suppression subtrative hybridization cDNAlibrary of Puccinellia tenuiflora roots grown on saline-alkali land, a total of94up-regulated unigenes was screened by cDNA macroarray hybridization. Blastx analysisshowed that70unigenes had high homology with the known genes, and the other24unigenes had no homology with the sequence in the database,which could beconsidered as as new genes.2. The up-regulated genes screened by macroarray hybridization were classfiedinto17founctional categories according to Arabidopsis thaliana MIPS standard, amongthem, the subcellular localization class accounts for the largest share, at18.26%,followed by protein class, at13.91%, function unknown class also accounted for a largepart of6.09%. The screened unigenes from Puccinellia tenuiflora roots involves indifferent biological process including energy metabolism, carbohydrate metabolism,protein synthesis and degradation, transcription regulation and signal transduction, etc.3. Macroarray hybridization screened out some unigenes closely related to plantsalt resistance, such as genes encoding universal stress protein, zinc finger protein,mitochondrial protein, glyceraldehyde3-phosphate dehydrogenase2, H+-ATPase,cytatin, light-induced protein1-like protein, cysteine protease, vacuolar processingenzyme, maturase K, and elongation factor. These genes provided important clues forrevealing molecular mechanism of plant salt tolerance and isolating salt-tolerant genesfrom Puccinellia tenuiflora. 4. RT-PCR technology was performed to analyze the expression characteristics ofgenes including Leg, Cytatin, HATPG, GAPDH2, Ring, WD40gene under both of150mM and200mM Na₂CO₃stressed conditions, seperately, the results showed that thesegenes were all induced by Na₂CO₃stresss, they might have important roles and behelpful for Puccinellia tenuiflora roots in defense of the adverse saline-alkali soilconditon.