Molecular and bioinformatics analysis of cDNA's selected from stem suppression subtractive hybridization library of Hordeum spontaneum C. Koch grown under zinc deficient conditions

Zinc (Zn) is an important micronutrient for all organisms and an essential component of over 1000 enzymes and proteins. Zinc plays a fundamental roles in protein, carbohydrate and lipid metabolisms, replication, transcription and translation and for structural and functional integrity of biomembranes.Zinc-deficiency causes many symptoms in all living things. In plants grown under low zinc conditions the general symptoms are stunting of shoot, curling and rolling of young leaves, death of leaf tips and chlorosis. In mammals low Zn in nutrition causes anemia, disorder of immunity and growth problems. Therefore, all organisms have evolved some mechanisms that tightly control zinc levels and respond to unexpected deficiency or excess of zinc in their cells.The crop plants that are resistant to zinc-deficiency should be cultivated to eliminate Zn-deficiency as a worldwide health problem. There are genotypic variations in zinc-uptake efficiency between plants species. These variations cause differential expression of genes encoding proteins used in uptake, transport and storage of zinc. In our research group Hordeum spontaneum C. Koch which is well adapted to Turkey?s zinc-deficient soils was used for high throughput identification of genes that are expressed upon zinc-deficiency. In our previous projects three different SSH library, including about total 5808 clones, were constructed using RNAs extracted from leaf, stem and root of Hordeum spontaneum C. Koch which was grown on zinc deficient medium. Sequence and bioinformatics analysis of 960 leaf and 384 root clones were completed.In this study we aimed isolation and identification of cDNAs that are found in stem SSH library. For this purpose 672 clones were selected randomly and sequenced. Raw sequence data was manipulated using some bioinformatics tools. Firstly low quality sequences, vector fragments and nonrecombinant clones were removed using Phred and Crossmatch programmes which are used with Cygwin software. Then contigs and singlets were generated using both CAP3 and Phrap programmes and results were compared.Sequences of contigs and singlets were subjected to nucleotid and protein BLAST. The sequences showed significant similarity to a variety of genes that were implicated to have roles in metabolism, defence and stress responses, structure and cellular organisation, transcription, translation, signal transduction, and apoptosis. Some of these genes are well characterised in terms of structure and function. These include Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase (RuBisCO), Lipid Transfer Protein, BLT4 protein, Chitinases, Actin, Pathogen-Related Protein, UDP-Glucose Pyrophosphorylase, Papain-like Sistein proteinase and CPN60 protein encoding genes.