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Characterizing the stress/defense transcriptome of Arabidopsis.

Mahalingam R, Gomez-Buitrago A, Eckardt N, Shah N, Guevara-Garcia A, Day P, Raina R, Fedoroff NV - Genome Biol. (2003)

Bottom Line: Several stress-responsive cis-elements showed a statistically significant over-representation in the promoters of the genes in the stress cDNA collection.These include W- and G-boxes, the SA-inducible element, the abscisic acid response element and the TGA motif.This set of stress-, pathogen- and hormone-modulated genes is an important resource for understanding the genetic interactions underlying stress signaling and responses and may contribute to the characterization of the stress transcriptome through the construction of standardized specialized arrays.

View Article: PubMed Central - HTML - PubMed

Affiliation: Life Sciences Consortium, Pennsylvania State University, State College, PA 16802, USA.

ABSTRACT

Background: To understand the gene networks that underlie plant stress and defense responses, it is necessary to identify and characterize the genes that respond both initially and as the physiological response to the stress or pathogen develops. We used PCR-based suppression subtractive hybridization to identify Arabidopsis genes that are differentially expressed in response to ozone, bacterial and oomycete pathogens and the signaling molecules salicylic acid (SA) and jasmonic acid.

Results: We identified a total of 1,058 differentially expressed genes from eight stress cDNA libraries. Digital northern analysis revealed that 55% of the stress-inducible genes are rarely transcribed in unstressed plants and 17% of them were not previously represented in Arabidopsis expressed sequence tag databases. More than two-thirds of the genes in the stress cDNA collection have not been identified in previous studies as stress/defense response genes. Several stress-responsive cis-elements showed a statistically significant over-representation in the promoters of the genes in the stress cDNA collection. These include W- and G-boxes, the SA-inducible element, the abscisic acid response element and the TGA motif.

Conclusions: The stress cDNA collection comprises a broad repertoire of stress-responsive genes encoding proteins that are involved in both the initial and subsequent stages of the physiological response to abiotic stress and pathogens. This set of stress-, pathogen- and hormone-modulated genes is an important resource for understanding the genetic interactions underlying stress signaling and responses and may contribute to the characterization of the stress transcriptome through the construction of standardized specialized arrays.

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Differential screening of clones from the stress libraries generated using SSH. Subtracted cDNA fragments obtained by the SSH procedure were cloned (see Materials and methods for details) and maintained as bacterial cultures in 96-well plates for each library. Quadruplicate colony dot blots were prepared and the membranes hybridized with labeled unsubtracted cDNA probes derived from (a) the driver, (b) the unsubtracted cDNA probes from the tester, (c) the forward subtracted cDNAs or (d) the reverse-subtracted cDNA.
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Figure 2: Differential screening of clones from the stress libraries generated using SSH. Subtracted cDNA fragments obtained by the SSH procedure were cloned (see Materials and methods for details) and maintained as bacterial cultures in 96-well plates for each library. Quadruplicate colony dot blots were prepared and the membranes hybridized with labeled unsubtracted cDNA probes derived from (a) the driver, (b) the unsubtracted cDNA probes from the tester, (c) the forward subtracted cDNAs or (d) the reverse-subtracted cDNA.

Mentions: One of the main advantages of SSH is that it normalizes the cDNA abundance so that cDNAs encoded by genes that are expressed infrequently, but nonetheless differentially, can be identified readily [33]. The efficiency of normalization is illustrated in Figure 2. The more uniform distribution of hybridization intensities obtained using the subtracted cDNA probe (Figure 2c) reflects the equalization in the concentrations of individual species present at markedly different concentrations in the initial unsubtracted cDNA populations (Figure 2b).


Characterizing the stress/defense transcriptome of Arabidopsis.

Mahalingam R, Gomez-Buitrago A, Eckardt N, Shah N, Guevara-Garcia A, Day P, Raina R, Fedoroff NV - Genome Biol. (2003)

Differential screening of clones from the stress libraries generated using SSH. Subtracted cDNA fragments obtained by the SSH procedure were cloned (see Materials and methods for details) and maintained as bacterial cultures in 96-well plates for each library. Quadruplicate colony dot blots were prepared and the membranes hybridized with labeled unsubtracted cDNA probes derived from (a) the driver, (b) the unsubtracted cDNA probes from the tester, (c) the forward subtracted cDNAs or (d) the reverse-subtracted cDNA.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC153460&req=5

Figure 2: Differential screening of clones from the stress libraries generated using SSH. Subtracted cDNA fragments obtained by the SSH procedure were cloned (see Materials and methods for details) and maintained as bacterial cultures in 96-well plates for each library. Quadruplicate colony dot blots were prepared and the membranes hybridized with labeled unsubtracted cDNA probes derived from (a) the driver, (b) the unsubtracted cDNA probes from the tester, (c) the forward subtracted cDNAs or (d) the reverse-subtracted cDNA.
Mentions: One of the main advantages of SSH is that it normalizes the cDNA abundance so that cDNAs encoded by genes that are expressed infrequently, but nonetheless differentially, can be identified readily [33]. The efficiency of normalization is illustrated in Figure 2. The more uniform distribution of hybridization intensities obtained using the subtracted cDNA probe (Figure 2c) reflects the equalization in the concentrations of individual species present at markedly different concentrations in the initial unsubtracted cDNA populations (Figure 2b).

Bottom Line: Several stress-responsive cis-elements showed a statistically significant over-representation in the promoters of the genes in the stress cDNA collection.These include W- and G-boxes, the SA-inducible element, the abscisic acid response element and the TGA motif.This set of stress-, pathogen- and hormone-modulated genes is an important resource for understanding the genetic interactions underlying stress signaling and responses and may contribute to the characterization of the stress transcriptome through the construction of standardized specialized arrays.

View Article: PubMed Central - HTML - PubMed

Affiliation: Life Sciences Consortium, Pennsylvania State University, State College, PA 16802, USA.

ABSTRACT

Background: To understand the gene networks that underlie plant stress and defense responses, it is necessary to identify and characterize the genes that respond both initially and as the physiological response to the stress or pathogen develops. We used PCR-based suppression subtractive hybridization to identify Arabidopsis genes that are differentially expressed in response to ozone, bacterial and oomycete pathogens and the signaling molecules salicylic acid (SA) and jasmonic acid.

Results: We identified a total of 1,058 differentially expressed genes from eight stress cDNA libraries. Digital northern analysis revealed that 55% of the stress-inducible genes are rarely transcribed in unstressed plants and 17% of them were not previously represented in Arabidopsis expressed sequence tag databases. More than two-thirds of the genes in the stress cDNA collection have not been identified in previous studies as stress/defense response genes. Several stress-responsive cis-elements showed a statistically significant over-representation in the promoters of the genes in the stress cDNA collection. These include W- and G-boxes, the SA-inducible element, the abscisic acid response element and the TGA motif.

Conclusions: The stress cDNA collection comprises a broad repertoire of stress-responsive genes encoding proteins that are involved in both the initial and subsequent stages of the physiological response to abiotic stress and pathogens. This set of stress-, pathogen- and hormone-modulated genes is an important resource for understanding the genetic interactions underlying stress signaling and responses and may contribute to the characterization of the stress transcriptome through the construction of standardized specialized arrays.

Show MeSH
Related in: MedlinePlus