Expression analysis of proliferative phase Arabidopsis thaliana endosperm

Laser capture microdissection (LCM) provides a useful method for isolating specific cells or tissues from biological samples. Here, we adapted microdissection protocols to allow high-resolution transcript analysis of different tissues from developing Arabidopsis seed. However, to obtain enough RNA for microarray analyses it was necessary to amplify the RNA. Microarray analyses, using endosperm derived RNA amplified by two-round IVT, reproducibly identified endosperm enriched marker genes. Keywords: LCM Endosperm Arabidopsis We carried out four hybridisations from three replicate endosperm samples microdissected from sections of 4 DAP siliques. We hybridised the endosperm samples against a universal control obtained from the tissues remaining post-microdissection scraped from one of the sample slides. This approach was used to minimise alterations in the measured differential expression due to possible differences in RNA quality between LCM prepared samples and conventional samples. Endosperm rep1 was labelled with Cy3, rep3 with Cy5 and rep2 was hybridised twice as dye swaps ie once labelled with Cy3 and once with Cy5.