Arabidopsis has emerged as an important model system for molecular plant biology. The extensive resources available for Arabidopsis make it an attractive system to study the molecular mechanisms involved in early seed development. During the early stages of seed development Arabidopsis endosperm is syncytial and proliferates rapidly through repeated rounds of mitosis without cytokinesis. This stage of endosperm development is both important in determining final seed size and is a model for studying various aspects of cellular and molecular biology, such as the cell cycle and genomic imprinting. However, the small size of Arabidopsis seed, the syncytial nature of the proliferative endosperm, and the surrounding maternal tissues make high throughput molecular analysis of the early endosperm technically difficult.
To get around this we used laser capture microdissection to enable transcript analysis of the early proliferative endosperm of Arabidopsis at 4 days after pollination (DAP). Microarray results identified several thousand genes with endosperm expression, including many that were endosperm preferred. A number of genes were validated by relative quantification PCR and were consistent with the findings of the microarray. Meta analysis of the endosperm transcriptome revealed a developmental program dominated by mitosis and under the influence of several phytohormones, predominated by cytokinin signaling.
The list of endosperm-preferred genes included all characterised imprinted genes in Arabidopsis. Imprinting is an epigenetic phenomenon by which genes are expressed predominantly from either their paternal or their maternal allele and very few imprinted genes have been identified in plants. The mono-allelic expression of the characterised imprinted genes appears to be limited to the endosperm where they provide important regulatory controls for seed development via direct effects on endosperm development. Genes from the endosperm-preferred list were screened for mono-allelic expression using sequence polymorphisms between the Colombia and Landsberg erecta ecotypes. We generated PCR products that spanned the polymorphisms of 67 genes from template obtained by laser capture of endosperm tissue from hybrid seed. Sequence analysis revealed three genes which gave strong allelic bias toward the maternal allele (At2g32460, At1g55550 and At2g21420) and one biased for the paternal allele (At1g47840).
In summary, laser capture microdissection has enabled high-resolution transcript analysis of the proliferative stage of Arabidopsis endosperm development. The data generated provides a useful resource providing novel insight into early seed development, facilitating both identification of endosperm expressed and novel imprinted genes.
| Identifer | oai:union.ndltd.org:ADTP/197587 |
| Date | January 2008 |
| Creators | Day, Robert Charles, n/a |
| Publisher | University of Otago. Department of Biochemistry |
| Source Sets | Australiasian Digital Theses Program |
| Language | English |
| Detected Language | English |
| Rights | http://policy01.otago.ac.nz/policies/FMPro?-db=policies.fm&-format=viewpolicy.html&-lay=viewpolicy&-sortfield=Title&Type=Academic&-recid=33025&-find), Copyright Robert Charles Day |
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