Plant & Animal Genome IX Conference Plant & Animal Genome IX Conference
Poster: Sequencing & EST
P01_57.html
This paper reports the development of a differential display technique that enables virtually every transcript in a tissue of interest to be assayed systematically. The technique uses a novel and simple approach to amplify subsets of cDNA fragments containing a 3' poly-A tract. We found that by resolving fragments on non-denaturing polyacrylamide gels, under conditions that favour heteroduplex analysis, allelic single nucleotide polymorphisms were detected. This enabled us to identify not only genic but also allelic differences in gene expression between tissues of interest. To increase the likelihood of detecting both abundantly expressed and rare mRNA transcripts on low-resolution gels, the cDNA population was divided into 2240 pools. However by using higher resolution gels the number of cDNA pools could potentially be reduced to 144. Using developing xylem tissue from Pinus radiata, we detected an average of 13 cDNA fragments per primer pair, and estimate that approximately 29,000 genes or alleles could be assayed.
In pines, wood properties vary dramatically with both cambial age and genotype. To identify genes that affect wood properties in Pinus radiata, developing xylem tissue was collected from five 24 year-old trees along a base-to-crown cambial-age gradient. Multiple wood property and cellular traits were measured from the sampled sections and these were used to define juvenile and mature wood-forming tissue. By using the differential display approach in a single tree, we could detect an average of four differentially expressed fragments per primer pair between juvenile and mature wood-forming tissue, allowing us to potentially isolate ~9000 differentially expressed genes and/or alleles from these tissues. The latest results will be presented.