Why choose SAGE?
A recent SAGE study of the human transcriptome (3.5 million transcripts from 19 tissues Velculescu 1999) revealed expression of 43,000 different genes in a single cell type with expression levels ranging from 0.3 to 9,417 transcript copies per cell. 83% of transcripts were present at levels as low as one copy per cell. The 633 most highly expressed genes accounted for 45% of the cellular mRNA mass fraction. Most unique transcripts were expressed at low levels (<5 copies per cell) with just under 25% of the mRNA mass of the cell comprising 94% of the unique transcripts expressed, as previously shown by reassociation kinetic studies. Only 52% of these low copy transcripts matched expressed sequences (mRNAs and ESTs) in GenBank/EMBL. Similar data for transcript abundance in an analysis of the mouse brain transcriptome (150,000 tags representing 42,738 unique transcripts) has recently been shown Chrast 2000. However, the 42,738 unique tags matched only approximately 4,000 known genes (there are ~6,000 mouse mRNA sequences in GenBank) and approximately 10,000 EST clusters of unknown function (from a total of ~70,000 mouse UniGene clusters), whereas the remaining transcript tags (76%), mainly for genes with low expression levels, had no match in the public databases Chrast 2000. The SAGE tag sequence for an unknown gene is sufficient information to generate longer cDNA fragments for gene identification Polyak 1997 and this process will be further facilitated as genome sequencing nears completion. Many genes not currently present in public databases will be predicted from accumulating genomic sequence and corresponding cDNAs will eventually be arrayed. However it is questionable whether available, or even developing microarray technologies will have the sensitivity to detect and quantitate the preponderance of low abundance transcripts given hybridization kinetic limitations. The power of SAGE for gene discovery, combined with the sensitivity of an in-depth SAGE analysis, as exemplified by the human and mouse studies described above and enabled by facilities such as those of the AGRF, will be exploited by the MBGP.
Last modified on the 18th November 2002.
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