Progress 03/01/06 to 06/30/09
Outputs
OUTPUTS: This goal of this project was to study allelic variation in DNA and RNA during endosperm development. To accomplish this goal we have used microarrays to profile gene expression levels in developing endosperm tissue. We harvested tissue at 13 and 19 days after pollination from B73, Mo17, B73xMo17 and Mo17xB73 ears. RNA isolated from this tissue was hybridized to Affymetrix microarrays. The data has been publicly released through the NCBI GEO website. We also developed a set of quantitative allele-specific SNP assays using the MassArray technology. These ~90 assays were used to compare the relative levels of the maternal and paternal alleles in DNA isolated from a series of time points. In addition, these same assays were used to compare transcript levels of the maternal and paternal alleles. This data set was publicly released as supplementary material along with a publication at Plant Physiology. PARTICIPANTS: Robert Stupar work as a post-doctoral researcher on this project. In addition, we are currently partnering with David Baulcombe to perform follow-up research that might further explain our findings. TARGET AUDIENCES: The target audience for this project is researchers that are studying seed phenotypes. Importantly, we are trying to understand genetic mechanisms that control phenotype and gene expression in endosperm tissue. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
We did not find any major differences in the level of the maternal and paternal alleles during development of the maize endosperm. However, we have made significant gains towards understanding differences in transcript levels during endosperm development. Our results from this project have been recently published at Plant Physiology. First, we found that the endosperm displays higher levels of non-additive expression that vegetative tissues. This includes a large number of genes that exhibit maternal-like or paternal-like expression patterns in the hybrid genotypes. Second, we demonstrated that a substantial proportion (~10%) of genes tested exhibit differential imprinting. This was the first application of a highly quantitative to study potential imprinting at a large number of genes. We found evidence of subtle imprinting at a large proportion of the genes. The project resources allowed us to generate and analyze this unique dataset. This project will likely have impacts towards understanding unique aspects of gene regulation that occur in endosperm tissue. We have also learned of exciting new data on small RNA production during seed development and are now collaborating with David Baulcombe's lab to determine whether the non-additive expression in endosperm may be due to small RNAs.
Publications
- No publications reported this period
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Progress 01/01/08 to 12/31/08
Outputs
OUTPUTS: This goal of this project was to study allelic variation in DNA and RNA during endosperm development. To accomplish this goal we have used microarrays to profile gene expression levels in developing endosperm tissue. We harvested tissue at 13 and 19 days after pollination from B73, Mo17, B73xMo17 and Mo17xB73 ears. RNA isolated from this tissue was hybridized to Affymetrix microarrays. The data has been publicly released through the NCBI GEO website. We also developed a set of quantitative allele-specific SNP assays using the MassArray technology. These ~90 assays were used to compare the relative levels of the maternal and paternal alleles in DNA isolated from a series of time points. In addition, these same assays were used to compare transcript levels of the maternal and paternal alleles. This data set was publicly released as supplementary material along with a publication at Plant Physiology. PARTICIPANTS: Robert Stupar work as a post-doctoral researcher on this project. In addition, we are currently partnering with David Baulcombe to perform follow-up research that might further explain our findings. TARGET AUDIENCES: The target audience for this project is researchers that are studying seed phenotypes. Importantly, we are trying to understand genetic mechanisms that control phenotype and gene expression in endosperm tissue. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
We did not find any major differences in the level of the maternal and paternal alleles during development of the maize endosperm. However, we have made significant gains towards understanding differences in transcript levels during endosperm development. Our results from this project have been recently published at Plant Physiology. First, we found that the endosperm displays higher levels of non-additive expression that vegetative tissues. This includes a large number of genes that exhibit maternal-like or paternal-like expression patterns in the hybrid genotypes. Second, we demonstrated that a substantial proportion (~10%) of genes tested exhibit differential imprinting. This was the first application of a highly quantitative to study potential imprinting at a large number of genes. We found evidence of subtle imprinting at a large proportion of the genes. The project resources allowed us to generate and analyze this unique dataset. This project will likely have impacts towards understanding unique aspects of gene regulation that occur in endosperm tissue. We have also learned of exciting new data on small RNA production during seed development and are now collaborating with David Baulcombe's lab to determine whether the non-additive expression in endosperm may be due to small RNAs.
Publications
- Stupar RM, Hermanson PJ, Springer NM. 2007. Nonadditive expression and parent-of-origin effects identified by microarray and allele-specific expression profiling of maize endosperm. Plant Physiol. 145(2):411-425.
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Progress 01/01/07 to 12/31/07
Outputs
This goal of this project was to study allelic variation in DNA and RNA during endosperm development. To accomplish this goal we have used microarrays to profile gene expression levels in developing endosperm tissue. We harvested tissue at 13 and 19 days after pollination from B73, Mo17, B73xMo17 and Mo17xB73 ears. RNA isolated from this tissue was hybridized to Affymetrix microarrays. The data has been publicly released through the NCBI GEO website. We also developed a set of quantitative allele-specific SNP assays using the MassArray technology. These ~90 assays were used to compare the relative levels of the maternal and paternal alleles in DNA isolated from a series of time points. In addition, these same assays were used to compare transcript levels of the maternal and paternal alleles. This data set was publicly released as supplementary material along with a publication at Plant Physiology.
Impacts
We did not find any major differences in the level of the maternal and paternal alleles during development of the maize endosperm. However, we have made significant gains towards understanding differences in transcript levels during endosperm development. Our results from this project have been recentl published at Plant Physiology. First, we found that the endosperm displays higher levels of non-additive expression that vegetative tissues. This includes a large number of genes that exhibit maternal-like or paternal-like expression patterns in the hybrid genotypes. Second, we demonstrated that a substantial proportion (~10%) of genes tested exhibit differentil imprinting. This was the first application of a highly quantitative to study potential imprinting at a large number of genes. We found evidence of subtle imprinting at a large proportion of the genes. The project resources allowed us to generate and analyze this unique dataset. This project will likely have
impacts towards understanding unique aspects of gene regulation that occur in endosperm tissue.
Publications
- Stupar RM, Hermanson PJ, Springer NM. Nonadditive expression and parent-of-origin effects identified by microarray and allele-specific expression profiling of maize endosperm. Plant Physiol. 2007 Oct;145(2):411-
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Progress 03/01/06 to 12/31/06
Outputs
Significant progress has been made towards the completion of our objectives during the past year. Our project involved work focused on two specific aims. The first aim involves monitoring the relative presence of the maternal and paternal allele during endosperm development. The second aim involves studying the relative expression of the maternal and paternal allele during endosperm development. A large proportion of the world's food supply is provided by the endosperm tissue of grains. However, we have a limited understanding of the regulatory processes that occur in this highly specialized tissue. A brief update will be provided for each of the two aims. For the first aim we are using two different approaches to study the relative amount of the maternal and paternal alleles. One approach is based on Southern blot analysis while the second approach utilizes a quantitative SNP-based assay. We collected endosperm tissue from reciprocal crosses of the maize inbreds B73
and Mo17 at 12, 14, 16, 18 and 20 days after pollination. The relative amount of the maternal and paternal alleles was then tested using Southern blot analysis (for three genes) or a quantitative SNP assay (for 67 genes). The analysis would detect whether certain genes showed altered maternal:paternal ratios as a result of differential amplification or degradation. To date, we have not observed any evidence of altered maternal:paternal ratios. However, we are performing a second study that includes later developmental time points. The second aim involves studies of gene expression in maize endosperms. We have studied gene expression patterns in 13 and 19 day after pollination maize endosperms using Affymetrix microarrays. The results were analyzed to study whether genes display dosage-dependent expression patterns. Surprisingly, we found many examples of non-dosage-dependent gene expression in the endosperm. We proceeded to test the allele-specific expression levels for a set of about
130 genes. While most genes displayed the expected expression patterns we did identify 6 genes that display a paternal bias and 3 genes with a maternal bias in expression level at both 13 and 19 days after pollination. These results are the basis for a manuscript that is in preparation.
Impacts
It could be argued that the endosperm is one of the most important plant tissues as a large percentage of human food is composed of plant endosperm. However, we still have a limited understanding of the dynamics of DNA replication and gene expression in the endosperm. This project aims to provide a more detailed understanding of the factors that influence the ratio of the maternal:paternal allele during endosperm development. In addition, we are documenting the relative gene expression of the two alleles in the developing endosperm tissue in order to provide a better understanding of the rate of parental bias and imprinting in controlling gene expression in the endosperm.
Publications
- No publications reported this period
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