TRANSCRIPTOME 2002: From Functional Genomics to Systems
Differential Gene Expression in Leishmania Major Friedlin
A. Saxena, S.Yan, L. Worthey, A. Leland, PJ Myler, K. Stuart, Seattle Biomedical Research Institute, WA
Leishmania, a flagellated protozoan is responsible for a wide spectrum of human diseases. The sequencing of Leishmania genome is currently underway. A large number of genes have been identified and a small proportion of these have been ascribed functions. However, it is essential to carry out functional studies in order to utilize the mass of data generated by the sequencing project. Using DNA microarrays it is possible to study the expression of thousands of genes at the same time. Microarrays, containing PCR amplified DNA from a random amplified genomic library of L. major Friedlin (Ref: Akopyants et al, MBP 113 (2001)), were hybridized with fluorescent probes made from procyclic and metacyclic RNA. The fluorescent signal showed an excellent level of signal to noise ratio. The data was normalised for background and probe intensity. The relative abundance of RNA for each spot was calculated. We observed statistically significant increase in signal intensity (1 to 5-folds, p<0.01) in 5% of DNAs with the metacyclics probe. Meanwhile in procyclics, 1.5 % of DNAs showed 1 to 3 folds increase in signal intensity. We also carried out flip-dye experiments to take into account the inherent ability of some species of RNA to bind to a particular dye. Northern blots were used to corroborate our results. We have identified several of genes up-regulated in both procyclics and metacyclics.
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