Salinity is one of the major constraints for crop yield because it limits plant growth and reduces both the crop yield and the use of agricultural land. Increased salt tolerance requires new genetic sources of salt tolerance, and more efficient techniques for identifying salt-tolerant germplasm.
The DD-RT-PCR technique (Differential Display-Reverse Transcription- Polymerase Chain Reaction) is one of these techniques, which are able to compare and identify changes in gene expression at the mRNA and the cDNA levels between any pair of contrasting genotypes. It was performed using mRNA extracted from the aerial part of two contrasting Tunisian barley genotypes (Sabra: tolerant and kelibia: sensitive accessions) subjected or not to salt stress.
In this study, we have used this technique (RT-PCR) to identify cDNAs corresponding to transcripts up- or down-regulated by salt stress in barley. Within 18 primer combinations (3 Oligod(T) x 6 arbitrary primers), we have identified a total of 58 differential display products which are over-expressed or disappeared in stressed samples indicating a qualitative and quantitative difference in the gene expression.
***The up-expressed fragment were eluted and sequenced at the Pasteur Institute of Tunis and then compared to those of the bank data (Genbank Barley) to determine sequences having an optimal alignment with the query.
The result was the identification of many salt-responsive transcripts fragments corresponding to hypothetic proteins (T17F15.140-like and Mei2-like), to some proteins involved in oxidative and heat stress (GAD1= Glutamic Acid Decarboxylase) or to proteins involved in the resistance to pathogens (ß-1,3-glucanase) and to anionic flux resistance. But the most important finding is the identification of genes encoding the Na+/H+ antiporter which sequestrate sodium into vacuole in the tolerant barley genotype.
These two last transcripts encoding the sequestration of Na could be used as markers for selecting salt tolerant genotypes in the program of varietals improvement to salt stress tolerance.