Location: Plant Science ResearchTitle: Transcript Profiling of Common Bean (Phaseolus vulgaris L.) Using the GeneChip(R) Soybean Genome Array: Optimizing Analysis by Masking Biased Probes Author
Submitted to: Biomed Central (BMC) Plant Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/7/2010
Publication Date: 5/7/2010
Publication URL: http://hdl.handle.net/10113/46583
Citation: Yang, S.H., Valdes-Lopez, O., Xu, W.W., Bucciarelli, B., Gronwald, J.W., Hernandez, G., Vance, C.P. 2010. Transcript Profiling of Common Bean (Phaseolus vulgaris L.) Using the GeneChip(R) Soybean Genome Array: Optimizing Analysis by Masking Biased Probes. Biomed Central (BMC) Plant Biology. 10:85. Available: http://www.biomedcentral.com/1471-2229/10/85. Interpretive Summary: Common bean, an herbaceous annual legume, is the most important grain legume for direct human consumption comprising approximately 50% of the grain legumes consumed world-wide. Additionally, common bean is the primary source of dietary protein in many developing countries. Despite its economic and nutritional importance, there are limited genomic resources currently available for advancing its genetic, genomic, and molecular studies. Specifically, a common bean whole genome DNA microarray is not presently available. DNA microarrays are commonly used to measure changes in gene expression levels for thousands of genes simultaneously. Currently available however, is the soybean whole genome DNA microarray. Common bean and soybean are close relative species and share significant DNA sequence similarity suggesting that the soybean DNA microarray may be used to study common bean gene expression. However, it is generally known that using a DNA microarray designed for one species to measure gene expression in a closely related species results in a reduction in the detection of expressed genes. In this report, we have evaluated the effectiveness of using the soybean DNA microarray for gene expression studies in common bean. We applied a methodology that corrects for the reduced efficiency of detecting expressed common bean genes when using the soybean DNA microarray. We validated this methodology by comparing the accuracy of measuring gene expression in common bean before and after correcting for reduced detection efficiency. By applying this methodology, the number of expressed common bean genes detected using the soybean DNA microarray was increased by about 3-fold. Many of the common bean genes detected in this study control important agronomic traits. The new knowledge discovered in this study will advance the development of improved common bean varieties needed to meet the nutritional needs of humans worldwide.
Technical Abstract: Common bean (Phaseolus vulgaris) and soybean (Glycine max) both belong to the Phaseoleae tribe and share significant coding sequence homology. This suggests that the GeneChip(R) Soybean Genome Array (soybean GeneChip) may be used for gene expression studies using common bean. To evaluate the utility of the soybean GeneChip for transcript profiling of common bean, we hybridized cRNAs purified from nodule, leaf, and root of common bean and soybean in triplicate to the soybean GeneChip. Initial data analysis showed a decreased sensitivity and specificity in common bean cross-species hybridization (CSH) GeneChip data compared to that of soybean. We employed a method that masked putative probes targeting inter-species variable (ISV) regions between common bean and soybean. A masking signal intensity threshold was selected that optimized both sensitivity and specificity. After masking for ISV regions, the number of differentially expressed genes identified in common bean was increased by 2.8-fold reflecting increased sensitivity. Quantitative RT-PCR analysis of 20 randomly selected genes and purine-ureide pathway genes demonstrated an increased specificity after masking for ISV regions. We also evaluated masked probe frequency per probe set to gain insight into the sequence divergence pattern between common bean and soybean. The sequence divergence pattern analysis suggested that the genes for basic cellular functions and metabolism were highly conserved between soybean and common bean. Additionally, our results show that some classes of genes, particularly those associated with environmental adaptation, are highly divergent. The masked probe frequency data also suggested that the gene expression pattern between soybean and common bean is potentially more consistent among highly conserved genes compared to genes with higher sequence divergence. In conclusion, transcript profiling in common bean can be done using the soybean GeneChip. However, a significant decrease in sensitivity and specificity can be expected. Problems associated with CSH GeneChip hybridization can be mitigated by masking probes that target ISV regions. In addition to transcript profiling CSH of the GeneChip in combination with masking probes in the ISV regions can be used for comparative ecological and/or evolutionary genomics studies.