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MicroSAGE is highly representative and reproducible but reveals major differences in gene expression among samples obtained from similar tissues.

Blackshaw S, Kuo WP, Park PJ, Tsujikawa M, Gunnersen JM, Scott HS, Boon WM, Tan SS, Cepko CL - Genome Biol. (2003)

Bottom Line: Serial analysis of gene expression using small amounts of starting material (microSAGE) has not yet been conclusively shown to be representative, reproducible or accurate.We show that microSAGE is highly representative, reproducible and accurate, but that pronounced differences in gene expression are seen between tissue samples taken from different individuals.MicroSAGE is a reliable method of comprehensively profiling differences in gene expression among samples, but care should be taken in generalizing results obtained from libraries constructed from tissue obtained from different individuals and/or processed or stored differently.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Genetics, Harvard Medical School and Howard Hughes Medical Institute, 200 Longwood Avenue, Boston, MA 02115, USA.

ABSTRACT

Background: Serial analysis of gene expression using small amounts of starting material (microSAGE) has not yet been conclusively shown to be representative, reproducible or accurate.

Results: We show that microSAGE is highly representative, reproducible and accurate, but that pronounced differences in gene expression are seen between tissue samples taken from different individuals.

Conclusions: MicroSAGE is a reliable method of comprehensively profiling differences in gene expression among samples, but care should be taken in generalizing results obtained from libraries constructed from tissue obtained from different individuals and/or processed or stored differently.

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Related in: MedlinePlus

SAGE tag abundance levels for tags in the 41-year-old peripheral retina SAGE library (retina kept at room temperature 5 h post-enucleation) and the 44-year-old peripheral retina library (retina kept at room temperature 45 min post-enucleation) for tags that differed at p ≤ 0.005; 139 such tags are shown.
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Figure 6: SAGE tag abundance levels for tags in the 41-year-old peripheral retina SAGE library (retina kept at room temperature 5 h post-enucleation) and the 44-year-old peripheral retina library (retina kept at room temperature 45 min post-enucleation) for tags that differed at p ≤ 0.005; 139 such tags are shown.

Mentions: The correlation coefficient between tag abundance levels in the 44-year-old and 88-year-old retinal samples, which were both processed 45 minutes after enucleation, is substantially higher than the correlation coefficient between the retinal libraries from the 44-year-old and 41-year-old individuals. This is so despite the fact that the 44-year-old and 88-year-old individuals differ not only in age but also in sex. This could mean that a major portion of the observed variance might result from differential processing and/or storage of the tissue before mRNA preparation. However, comparison of expression of genes that differ by p ≤ 0.005 in the SAGE libraries from the peripheral retinas from the 44-year-old and the 41-year-old individuals indicates a roughly equal magnitude and direction of change of overall expression levels between the samples (Figure 6). Were the differences between the samples largely due to differential mRNA stability, one would expect to see a subset of mRNAs that were much reduced in expression in the 41-year-old sample, as that one spent a longer period of time between enucleation and RNA extraction. One might also expect to see a fraction of more stable transcripts showing a small, compensatory increase in relative frequency, but this is not the case. The most likely explanation then for the differences between the 41-year-old and 44-year-old samples is variation in gene expression.


MicroSAGE is highly representative and reproducible but reveals major differences in gene expression among samples obtained from similar tissues.

Blackshaw S, Kuo WP, Park PJ, Tsujikawa M, Gunnersen JM, Scott HS, Boon WM, Tan SS, Cepko CL - Genome Biol. (2003)

SAGE tag abundance levels for tags in the 41-year-old peripheral retina SAGE library (retina kept at room temperature 5 h post-enucleation) and the 44-year-old peripheral retina library (retina kept at room temperature 45 min post-enucleation) for tags that differed at p ≤ 0.005; 139 such tags are shown.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC153457&req=5

Figure 6: SAGE tag abundance levels for tags in the 41-year-old peripheral retina SAGE library (retina kept at room temperature 5 h post-enucleation) and the 44-year-old peripheral retina library (retina kept at room temperature 45 min post-enucleation) for tags that differed at p ≤ 0.005; 139 such tags are shown.
Mentions: The correlation coefficient between tag abundance levels in the 44-year-old and 88-year-old retinal samples, which were both processed 45 minutes after enucleation, is substantially higher than the correlation coefficient between the retinal libraries from the 44-year-old and 41-year-old individuals. This is so despite the fact that the 44-year-old and 88-year-old individuals differ not only in age but also in sex. This could mean that a major portion of the observed variance might result from differential processing and/or storage of the tissue before mRNA preparation. However, comparison of expression of genes that differ by p ≤ 0.005 in the SAGE libraries from the peripheral retinas from the 44-year-old and the 41-year-old individuals indicates a roughly equal magnitude and direction of change of overall expression levels between the samples (Figure 6). Were the differences between the samples largely due to differential mRNA stability, one would expect to see a subset of mRNAs that were much reduced in expression in the 41-year-old sample, as that one spent a longer period of time between enucleation and RNA extraction. One might also expect to see a fraction of more stable transcripts showing a small, compensatory increase in relative frequency, but this is not the case. The most likely explanation then for the differences between the 41-year-old and 44-year-old samples is variation in gene expression.

Bottom Line: Serial analysis of gene expression using small amounts of starting material (microSAGE) has not yet been conclusively shown to be representative, reproducible or accurate.We show that microSAGE is highly representative, reproducible and accurate, but that pronounced differences in gene expression are seen between tissue samples taken from different individuals.MicroSAGE is a reliable method of comprehensively profiling differences in gene expression among samples, but care should be taken in generalizing results obtained from libraries constructed from tissue obtained from different individuals and/or processed or stored differently.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Genetics, Harvard Medical School and Howard Hughes Medical Institute, 200 Longwood Avenue, Boston, MA 02115, USA.

ABSTRACT

Background: Serial analysis of gene expression using small amounts of starting material (microSAGE) has not yet been conclusively shown to be representative, reproducible or accurate.

Results: We show that microSAGE is highly representative, reproducible and accurate, but that pronounced differences in gene expression are seen between tissue samples taken from different individuals.

Conclusions: MicroSAGE is a reliable method of comprehensively profiling differences in gene expression among samples, but care should be taken in generalizing results obtained from libraries constructed from tissue obtained from different individuals and/or processed or stored differently.

Show MeSH
Related in: MedlinePlus