|Public on Apr 25, 2008
|Arabidopsis globular stage chalazal seed coat captured by LCM
|Siliques with length between 1.1 and 1.4 cm containing globular stage seeds were collected from 38-46 day old plants. Seeds were either dissected out of siliques or left in sub-divided siliques and fixed in 3:1 (v/v) ethanol to acetic acid and embedded in paraffin. Microdissection of compartments was carried out on 5 or 7 um sections using a Leica LMD6000 system. Compartments were only captured from seeds with globular embryos 24-32 um in diameter, as determined by a medial section through the embryo proper.
|Plants were grown in a Conviron chamber under continuous light with fluorescent lamps at 20°C and 50% - 70% relative humidity.
|Total RNA was extracted using the RNaqueous Micro-Kit (Ambion, Austin, TX) using a modified protocol incorporating an on-column DNase treatment (Qiagen RNase-Free DNase Set) during column washes.
|2 to 4 ng of total RNA was amplified using the WT-Ovation Pico RNA Amplification System (NuGEN, San Carlos, CA). cDNAs were fragmented and labeled with NuGEN FL-Ovation™ cDNA Biotin Module V2.
|Following fragmentation, five micrograms of cDNA was hybridized for 18 hr at 45°C with the GeneChip ATH1 Arabidopsis Genome Array. GeneChips were washed and stained using the EukGE-WS2V4_450 protocol in the Affymetrix Fluidics Station 450.
|GeneChips were scanned using the Affymetrix GeneChip Scanner 3000 7G System.
|Laser capture microdissected (LCM) chalazal seed coat at the globular stage, biological replicate 1
Chalazal seed coat collected from seeds containing globular embryos 24 to 32 um in diameter as determined by median section using the LEICA LMD6000 system for microdissection.
|The data were analyzed with the Affymetrix GeneChip Operating system 1.4 (GCOS) using default analysis settings and global scaling as a normalization method. The mean target intensity of each array was set to 500.
|Apr 25, 2008
|Last update date
|Aug 28, 2018
|John J Harada
|University of California, Davis
|One Shields Avenue
|Expression data from Arabidopsis Seed Compartments at the Globular Embryo Stage.
|Expression data from Arabidopsis Seed Compartments at 5 discrete stages of development