Comparing two microarray platforms for identifying mutated genes in barley (Hordeum vulgare L.).
We have previously described the evaluation of a cDNA microarray platform to identify and clone mutated barley (Hordeum vulgare L.) genes, using their transcriptionally defective mutant alleles (S. Zakhrabekova, C.G. Kannangara, D. von Wettstein, M. Hansson, A microarray approach for identification of mutated genes, Plant Physiol. Biochem. 40 (2002) 189-197). It was concluded that competitive hybridization between phenotypically similar mutants could specifically highlight an arrayed clone, corresponding to the mutated gene. In this study we evaluate whether the Affymetrix microarray platform can be used for the same purpose. The Affymetrix barley microarray contains a large number of clones (22,792 probe sets). In this and the previous study we used two barley mutant strains, xantha-h.57 and xantha-f.27, with known mutations in different subunit genes of the chlorophyll biosynthetic enzyme magnesium chelatase (EC 6.6.1.1). Mutant xantha-h.57 produces no Xantha-h mRNA whereas in xantha-f.27 the nonsense mutation in the last exon of the gene, results in nonsense-mediated decay of Xantha-f mRNA. We conclude that the Affymetrix platform meets our requirements and that our approach successfully highlighted the arrayed Xantha-h clone and that Xantha-f was among the top fourteen candidates.
Zakhrabekova
Shakhira
S
Department of Biochemistry, Lund University, Box 124, SE-22100 Lund, Sweden. shakhira.zakhrabekova@biochemistry.lu.se
Gough
Simon P
SP
Lundqvist
Udda
U
Hansson
Mats
M
D000483:Alleles; D002734:Chlorophyll; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D020869:Gene Expression Profiling; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D001467:Hordeum / Q000235:genetics*; D008190:Lyases / Q000378:metabolism; D008957:Models, Genetic; D009154:Mutation; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation*; D010940:Plant Proteins / Q000235:genetics*; D012313:RNA / Q000737:chemistry
D010940:Plant Proteins; D002734:Chlorophyll; D012313:RNA; D008190:Lyases; C086788:magnesium chelatase