African Journal of Biotechnology Vol. 2 (4), pp. 75-81, April 2003

Functional and comparative analysis of expressed sequences from Diuraphis noxia infested wheat obtained utilizing the conserved Nucleotide Binding Site

Lynelle Lacock, Chantal van Niekerk, Shilo Loots, Franco du Preez and Anna-Maria Botha*

Department of Genetics, Forestry and Agricultural Biotechnology Institute (FABI), University of Pretoria, Pretoria, ZA0002 South Africa

*Corresponding Author; E-mail: ambothao@postino.up.ac.za, tel: +27 12 420 3945, fax: +27 12 420 3947

Accepted 10 March 2003

Russian wheat aphid (Diuraphis noxia, Morvilko; RWA) is a major pest on wheat, barley and other triticale in South Africa.  Infestation by the RWA results in altered protein expression patterns, which is manifested as differential expression of gene sequences.  In the present study, Russian wheat aphid resistant (Tugela DN, Tugela*5/SA2199, Tugela*5/SA463, PI 137739, PI 262660, and PI 294994) and susceptible triticale (Tugela) were infested and cDNA synthesized.  A PCR based approach was utilized to amplify the nucleotide binding site conserved region to obtain expressed sequence tags (ESTs) with homology to resistance gene analogs (RGAs).  The approach proved highly feasible when the isolation of RGAs is the main objective, since 18% of all obtained ESTs showed significant hits with known RGAs, when translated into their corresponding amino acid sequences and searched against the nonredundant GenBank protein database using the BLASTX algorithm.

Key words:  Resistance gene analogs, degenerate PCR, nucleotide-binding site-leucine rich repeat resistance genes, Aegilops tauschii.