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A novel design of whole-genome microarray probes for Saccharomyces cerevisiae which minimizes cross-hybridization.

Talla E, Tekaia F, Brino L, Dujon B - BMC Genomics (2003)

Bottom Line: We present here a novel design of Saccharomyces cerevisiae microarrays based on a refined annotation of the genome and with the aim of reducing cross-hybridization between related sequences.The sequence of each gene was compared against the entire yeast genome and optimal sub-segments giving no predicted cross-hybridization were selected.The OliD program is available from authors.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institut Pasteur, Unité de Génétique Moléculaire des Levures (URA 2171 CNRS, UFR 927 Université PM Curie), 25 rue du Docteur Roux, F-75724 Paris cedex 15, France. etalla@pasteur.fr

ABSTRACT

Background: Numerous DNA microarray hybridization experiments have been performed in yeast over the last years using either synthetic oligonucleotides or PCR-amplified coding sequences as probes. The design and quality of the microarray probes are of critical importance for hybridization experiments as well as subsequent analysis of the data.

Results: We present here a novel design of Saccharomyces cerevisiae microarrays based on a refined annotation of the genome and with the aim of reducing cross-hybridization between related sequences. An effort was made to design probes of similar lengths, preferably located in the 3'-end of reading frames. The sequence of each gene was compared against the entire yeast genome and optimal sub-segments giving no predicted cross-hybridization were selected. A total of 5660 novel probes (more than 97% of the yeast genes) were designed. For the remaining 143 genes, cross-hybridization was unavoidable. Using a set of 18 deletant strains, we have experimentally validated our cross-hybridization procedure. Sensitivity, reproducibility and dynamic range of these new microarrays have been measured. Based on this experience, we have written a novel program to design long oligonucleotides for microarray hybridizations of complete genome sequences.

Conclusions: A validated procedure to predict cross-hybridization in microarray probe design was defined in this work. Subsequently, a novel Saccharomyces cerevisiae microarray (which minimizes cross-hybridization) was designed and constructed. Arrays are available at Eurogentec S. A. Finally, we propose a novel design program, OliD, which allows automatic oligonucleotide design for microarrays. The OliD program is available from authors.

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Schematic representation of the design procedure. The main scripts (written in perl and sh shell programming languages) used in this work are in brackets []. [Newsort] allows the rearrangement of the blast output table in a positional order along the query and subject sequences. [Verifalign] detects the different types of alignments (as described in Figure 1). [Verifbarres] and [Verifbar] allow the detection of potential cross-hybridization regions in Type2 and Type1 alignments, respectively. All scripts are available upon request.
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Figure 5: Schematic representation of the design procedure. The main scripts (written in perl and sh shell programming languages) used in this work are in brackets []. [Newsort] allows the rearrangement of the blast output table in a positional order along the query and subject sequences. [Verifalign] detects the different types of alignments (as described in Figure 1). [Verifbarres] and [Verifbar] allow the detection of potential cross-hybridization regions in Type2 and Type1 alignments, respectively. All scripts are available upon request.

Mentions: Our probe design involves 3 steps as illustrated by Figure 5.


A novel design of whole-genome microarray probes for Saccharomyces cerevisiae which minimizes cross-hybridization.

Talla E, Tekaia F, Brino L, Dujon B - BMC Genomics (2003)

Schematic representation of the design procedure. The main scripts (written in perl and sh shell programming languages) used in this work are in brackets []. [Newsort] allows the rearrangement of the blast output table in a positional order along the query and subject sequences. [Verifalign] detects the different types of alignments (as described in Figure 1). [Verifbarres] and [Verifbar] allow the detection of potential cross-hybridization regions in Type2 and Type1 alignments, respectively. All scripts are available upon request.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC239980&req=5

Figure 5: Schematic representation of the design procedure. The main scripts (written in perl and sh shell programming languages) used in this work are in brackets []. [Newsort] allows the rearrangement of the blast output table in a positional order along the query and subject sequences. [Verifalign] detects the different types of alignments (as described in Figure 1). [Verifbarres] and [Verifbar] allow the detection of potential cross-hybridization regions in Type2 and Type1 alignments, respectively. All scripts are available upon request.
Mentions: Our probe design involves 3 steps as illustrated by Figure 5.

Bottom Line: We present here a novel design of Saccharomyces cerevisiae microarrays based on a refined annotation of the genome and with the aim of reducing cross-hybridization between related sequences.The sequence of each gene was compared against the entire yeast genome and optimal sub-segments giving no predicted cross-hybridization were selected.The OliD program is available from authors.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institut Pasteur, Unité de Génétique Moléculaire des Levures (URA 2171 CNRS, UFR 927 Université PM Curie), 25 rue du Docteur Roux, F-75724 Paris cedex 15, France. etalla@pasteur.fr

ABSTRACT

Background: Numerous DNA microarray hybridization experiments have been performed in yeast over the last years using either synthetic oligonucleotides or PCR-amplified coding sequences as probes. The design and quality of the microarray probes are of critical importance for hybridization experiments as well as subsequent analysis of the data.

Results: We present here a novel design of Saccharomyces cerevisiae microarrays based on a refined annotation of the genome and with the aim of reducing cross-hybridization between related sequences. An effort was made to design probes of similar lengths, preferably located in the 3'-end of reading frames. The sequence of each gene was compared against the entire yeast genome and optimal sub-segments giving no predicted cross-hybridization were selected. A total of 5660 novel probes (more than 97% of the yeast genes) were designed. For the remaining 143 genes, cross-hybridization was unavoidable. Using a set of 18 deletant strains, we have experimentally validated our cross-hybridization procedure. Sensitivity, reproducibility and dynamic range of these new microarrays have been measured. Based on this experience, we have written a novel program to design long oligonucleotides for microarray hybridizations of complete genome sequences.

Conclusions: A validated procedure to predict cross-hybridization in microarray probe design was defined in this work. Subsequently, a novel Saccharomyces cerevisiae microarray (which minimizes cross-hybridization) was designed and constructed. Arrays are available at Eurogentec S. A. Finally, we propose a novel design program, OliD, which allows automatic oligonucleotide design for microarrays. The OliD program is available from authors.

Show MeSH