High-density cDNA filter analysis: a novel approach for large-scale, quantitative analysis of gene expression

Zhao, Nan; Hashida, Hideji; Takahashi, Nobuaki; Misumi, Yoshio; Sakaki, Yoshiyuki

doi:10.1016/0378-1119(95)00023-y

Cited by 111 publications

(40 citation statements)

References 15 publications

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“…Several studies have examined the compara-tive aspects of microarrays with standard methodologies including Northern/dot blotting (8) and quantitative PCR (9) and have demonstrated that expression of even rare gene transcripts (less than 0.1%) can be measured quantitatively (10). There has been little discussion, however, regarding the ability of arrays to measure small changes in gene expression and whether these approaches have dynamic range similar to standard approaches such as Northern blot analysis.…”

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confidence: 99%

Development of a Toxicological Gene Array and Quantitative Assessment of This Technology

Bartosiewicz

Trounstine²,

Barker³

et al. 2000

Archives of Biochemistry and Biophysics

113

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High-density arrays of DNA bound to solid substrates offer a powerful approach to identifying changes in gene expression in response to toxicants. While DNA arrays have been used to explore qualitative changes in gene regulation, less attention has focused on the quantitative aspects of this technology. Arrays containing expressed sequence tags for xenobiotic metabolizing enzymes, proteins associated with glutathione regulation, DNA repair enzymes, heat shock proteins, and housekeeping genes were used to examine gene expression in response to ␤-naphthoflavone (␤-NF). Upregulation of cytochrome P4501a1 (Cyp1a1) and 1a2 in mouse liver was maximal 8 h after ␤-NF administration. Significant upregulation of Cyp1a2 was noted at ␤-NF doses as low as 0.62 and 1.2 mg/kg when gene expression was measured by microarray or Northern blotting, respectively. Maximal Cyp1a2 induction is 5-fold by Northern analysis and 10-fold by microarray. Induction of Cyp1a1 was 15-and 20-fold by Northern and microarray analysis, respectively. The coefficient of variation for spot to spot and slide to slide comparisons was <15%; this variability was smaller than interanimal variability (18 -60%). Comparison of mRNA expression in control animals indicated that there are differences in labeling/detection associated with Cy3/Cy5 dyes; accordingly, experiments must include methods for establishing baseline signals for all genes. We conclude that the dynamic range and sensitivity of DNA microarrays on glass slides is comparable to Northern blotting analysis and that variability of the data introduced during spotting and hybridization is less than the interanimal variability.

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mentioning

confidence: 99%

Development of a Toxicological Gene Array and Quantitative Assessment of This Technology

Bartosiewicz

Trounstine²,

Barker³

et al. 2000

Archives of Biochemistry and Biophysics

113

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“…Subtractive cDNA cloning, differential display, and DNA microarrays have been used to analyze global gene expression under different growth and environmental conditions (22,40,43,46). These procedures are easier to perform in eukaryotes, due to the presence of a poly(A) tail on mRNA that permits easy separation from rRNA (3).…”

mentioning

confidence: 99%

Identification of Iron-Responsive, Differential Gene Expression in the CyanobacteriumSynechocystissp. Strain PCC 6803 with a Customized Amplification Library

Singh

Sherman

2000

J Bacteriol

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We describe the use of a method called differential expression using customized amplification library (DECAL) to study the global changes in gene expression in iron-deficient versus iron-reconstituting cells of Synechocystis sp. strain PCC 6803. We identified a number of genes, such as isiA, idiA, psbA, cpcG, and slr0374, whose expression either increased or decreased in response to iron availability. Further analysis led to the identification of additional genes related to those identified by DECAL (e.g., psbC, psbO, psaA, apcABC, cpcBAC1C2D, and nblA) that were differentially regulated by iron availability. Expression of cpcG, psbC, psbO, psaA, apcABC, and cpcBAC1C2D increased, whereas that of isiA, idiA, nblA, psbA, and slr0374 decreased, in iron-reconstituting cells. S1 nuclease protection studies showed that increased transcript levels of psbA in iron-deficient cells was due to the increased expression of both psbA2 and psbA3 genes, although the steadystate level of psbA2 remained higher than that of psbA3.

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“…In contrast to functional assays, the quantitative analysis of gene expression level lends itself to largescale implementation. Two main approaches have been proposed (1) ''analog'' methods based on hybridization to arrayed cDNA libraries (Lennon and Lehrach 1991;Gress et al 1992;Nguyen et al 1995;Schena et al 1995;Zhao et al 1995) or oligonucleotide ''chips'' (Fodor et al 1991;Southern et al 1992;Guo et al 1994;Matson et al 1995); and (2) ''digital'' methods, based on the generation of sequence tags. This paper focuses on the latter.…”

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confidence: 99%

The Significance of Digital Gene Expression Profiles

Audic

Claverie²

1997

Genome Res.

2,598

2,130

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Genes differentially expressed in different tissues, during development, or during specific pathologies are of foremost interest to both basic and pharmaceutical research. “Transcript profiles” or “digital Northerns” are generated routinely by partially sequencing thousands of randomly selected clones from relevant cDNA libraries. Differentially expressed genes can then be detected from variations in the counts of their cognate sequence tags. Here we present the first systematic study on the influence of random fluctuations and sampling size on the reliability of this kind of data. We establish a rigorous significance test and demonstrate its use on publicly available transcript profiles. The theory links the threshold of selection of putatively regulated genes (e.g., the number of pharmaceutical leads) to the fraction of false positive clones one is willing to risk. Our results delineate more precisely and extend the limits within which digital Northern data can be used.

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High-density cDNA filter analysis: a novel approach for large-scale, quantitative analysis of gene expression

Cited by 111 publications

References 15 publications

Development of a Toxicological Gene Array and Quantitative Assessment of This Technology

Development of a Toxicological Gene Array and Quantitative Assessment of This Technology

Identification of Iron-Responsive, Differential Gene Expression in the CyanobacteriumSynechocystissp. Strain PCC 6803 with a Customized Amplification Library

The Significance of Digital Gene Expression Profiles

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