Due to the increasing prevalence and severity of Al phytoxicity in certain regions of the South African sugar industry, a research programme has been initiated at SASEX to elucidate the molecular mechanisms by which sugarcane detects and responds to the metal. As part of this larger investigation, the current study aimed to assess the response of a reportedly Al tolerant cultivar, Saccharum spp. hybrid cv. N12, to phytotoxic levels of Al. Hydroponically-grown plants of this commercial genotype were used in Al inhibition studies, the results of which indicated that exposure of plants to 250µM Al for 24 hours resulted in maximum reduction of root elongation. Under these conditions, root growth was inhibited by approximately 36%, compared with only 4% for the 50µM Al treatment. Subsequently, this exposure regime was used to gather the terminal 5 to 10mm of root tips, the site of the primary Al lesion, of challenged and control, unchallenged plants for molecular analysis. Total RNA was extracted from the Al challenged and control root tips, from which mRNA was subsequently isolated, reverse transcribed and converted to double-stranded cDNA. The two populations of cDNA were reciprocally subtracted from each other and used to construct subtractive cDNA libraries in Lambda ZAP®II phages. Randomly selected clones, 576 representatives from each of the libraries, were screened using membrane-based array technology. Results indicated that only 33% (190) of the Al-treatment specific library cDNAs were found to be more highly expressed under conditions of Al stress than under control conditions. Of these potentially Al response-related cDNAs, 25 were sequenced and submitted to sequence databases for the assignment of putative identities. No genic sequences known to be directly associated with the Al stress response were identified, however, several were found to be related to pathogenesis or general stress pathways. Although further Northern hybridisation work is required to validate these results, they suggest that the induction of general stress response pathways may be involved in the aluminium stress response of this sugarcane cultivar. Such Al stress-related sequences could have applications in marker-assisted breeding programmes and as candidate genes for the genetic engineering of tolerant genotypes.

Thesis (M.Sc.)-University of Natal, Durban, 2002.