发信人: cyberpostdoc (cyberpostdoc), 信区: Biology
标 题: Re: Comment on the EST technology
发信站: BBS 未名空间站 (Sun Jun 26 23:59:56 2005), 转信

Your concern is exactly the key limitation of this technology. The advantage of EST is that it provides direct evidence of the expression of a gene. But several drawbacks I could think of:

1. mRNA abundance limitation: it is estimated that genes have an average expression level below 5% of the mRNA population is less likely to be detected by EST technology. Therefore, genes that are expressed at extreme low level won't be able to be detected by EST, so the 27K in this sense should be an lower estimate (if we only focus on this matter). Now, one of the other technique is more sensitive, it is called SAGE, for serial analysis of gene expression, where tags at 3'end of genes are collected and sequenced to represent mRNA expression.

2. development stage limitation: some of the genes in our genome may only express at certain development stage, therefore, if EST libraries cannot cover this development stage, both for technical and ethical reasons, there is no way these gene can be detected.

3. diseased stage limitation: on the same line of #2, but from an opposite point of view, some mRNA might be a result of diseased stage, not neccissary be a normally expressed gene. Or aberrent expression that are not constitute normal behavior of human genome, hence, enriched EST libraries from cancer cell lines (for example) will likely to result in over estimation of human transcriptome.

4. sequencing error: this could post extreme difficulties for bioinformatics to further map these ESTs back to genome to identify genes. Especially paralog genes.

Finally, beside gene discovery, EST can be used to study post transcriptional controls on a genomic level, which have been shown very successful, including alternative splicing and alternative polyadenylation. Pioneer of these include Michael Zhang at CSHL, Chris Burge at MIT, and Chris Lee at UC.

Hope these help!

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【 在 Marble (小石头哥哥) 的大作中提到: 】
: I was listening to a speech given by J. Craig Venter the other day. It was a
: very impressive one, though it was much more like elementary education to the
: general audience. One thing that really impressed me is the expressed sequence
: tags technology (ESTs), which was developed and applied by J. Craig Venter to
: discover a multitude of new genes and consequently integrated into the Human
: Genome Project. The explosion of discovering new genes with the help of
: mathmatics and computers was realized only with the immerge of the EST
: technology. Basically, the current estimated genes for human beings are about
: 26,000, and 23,000 for the Mus musculus. This number is pretty surprising to
: most biologists due to the large size of the genome, which means that almost
: ...................

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※ 修改:·cyberpostdoc 于 Jun 27 00:05:39 修改本文·[FROM: 67.82.]
※ 修改:·cyberpostdoc 於 Jun 28 21:27:47 2005 修改本文·[FROM: 67.82.198.137]
※ 修改:·cyberpostdoc 於 Jun 28 21:29:18 2005 修改本文·[FROM: 67.82.198.137]
※ 来源:·BBS 未名空间站 mitbbs.com·[FROM: 67.82.]