| A PCR based suppression subtractive hybridization (SSH) method was used to identify differentially expressed genes between control and transgenic aspen down-regulated in 4CL1 (4-coumarate:coenzyme A ligase) expression, and showing enhanced growth. A total of 11,308 expressed sequence tags (SSTs) representing 5,028 non-redundant transcripts encoding 4,224 unique proteins were obtained from shoot apex, young stem, young leaf, and root tip SSH libraries. Putative functions could be assigned to 60% of these transcripts. Sequence data was annotated and a database was created for storage and dissemination of sequence information. Efficient and automated procedure for data handling was facilitated by a series of PERL scripts. In this automated pipeline, the output from sequencing machines as chromatograms were subjected to base-calling, quality control, BLAST and annotation processes and finally to storage of this data in a database accessible through web interface. One of the interesting observations in the EST collection was the biased presence of homologs of transposable elements, especially copia elements, in transgenic tissues. In the absence of a fully sequenced Populus genome at the time this study was conducted, Arabidopsis genome was used to characterize the distribution, diversity, and expression of copia elements. Most of the elements are defective, reflecting ancient transposition activities. These elements are preferentially localized in gene-poor region around the centromere. The transcription of these elements appears to be tightly controlled. |
