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New Microarray Labeling Kit for Preparing cDNA Probes

added to the probe-labeling reaction correspond to mRNA of high, medium, and low abundances, respectively. The relative fluorescent intensities of the A. thaliana cDNA probes when hybridized to the corresponding A. thaliana target DNA on the Control Microarray can therefore be used to determine the sensitivity of the assay (Figure 1).

Fig.1

The 3X SSC negative controls can be used to determine the specificity of the assay because the labeled probe should not hybridize to the spotted 3X SSC.

RNA Quality

The quality of the RNA is critical to successful probe preparation. The presence of contaminants, such as carbohydrates, can increase background fluorescence following hybridization. The guanidinium isothiocyanate method used in the StrataPrep total RNA miniprep kit2 has been used successfully in this application.

The human -actin and additional target gene-labeled probes are generated from the researchers test or control mRNA in the probe-labeling reaction. The abundances of these mRNA will vary with each tissue. The fluorescent intensities of the -actin and other human gene cDNA probes when hybridized to the corresponding target DNA on the Control Microarray can therefore be used to determine the quality of the researchers mRNA.

Conclusions

Use Stratagenes Microarray Labeling Kit to prepare reliable cDNA probes for hybridization to gene microarrays. The Arabidopsis control mRNA and Control Microarray included in the kit allow problems associated with poor-quality mRNA to be identified prior to hybridizing the probe to an expensive microarray and to determine the sensitivity and specificity of the hybridization.

REFERENCES

  1. Schena, M., et al. (1995) Science 270: 467-470.

  2. '"/>



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