Analysis of Gene Expression during Brassica Seed Germination Using a Cross- Species Microarray Platform

We have developed an approach to facilitate the use of model organism microarrays in related, nonmodel organisms. We demonstrate the method by using Arabidopsis oligonucleotide microarrays to analyze gene expression in Brassica. Probes with low hybridization signals to Brassica genomic DNA were excluded from the transcriptional analysis of an Arabidopsis microarray at the software level, forming a virtual Brassica microarray. We then performed an experiment on transcriptional responses during seed germination in Brassica using 17 886 homologous probesets of the virtual array where Brassica mRNA hybridization was detected. We subjected seed to hydration or priming (a step to improve germination vigor) and subsequent heating and drying treatments (steps to prolong shelf life of dried seed). Exploration of the microarray results indicated two likely expression patterns shared by many genes. One class of transcripts was strongly, globally, and irreversibly downregulated by priming, while other transcripts were induced, but often reversibly. Legacy seed-storage protein messages were in the fi rst class, and many protein synthesis components and some resource mobilization enzymes were in the second. We were able to validate our results by confi rming transcriptional responses using reverse transcription polymerase chain reaction (RT–PCR). Brassica is a model system for the molecular processes during hydration of seeds. It has well-established, commercially useful seed treatment procedures, and Brassica species have signifi cant economic value as crops. Its close phylogenetic relationship to Arabidopsis thaliana makes it a clear potential benefi ciary of Arabidopsis functional genomics (Paterson et al., 2001). Caulifl ower (Brassica oleracea L. convar botrytis L. Alef. var. botrytis) is a vegetable crop for which the seed hydration treatment known as priming is well established and applied on a commercial scale. Priming of seeds is used to increase germination effi ciency. Under this treatment, seeds are partially hydrated and begin the processes of germination, but emergence does not occur, and the seeds are subsequently dried. Priming has several eff ects on the physiology of seeds, including developmental advancement, decreased membrane permeability, dormancy breaking, and the induction of DNA and protein synthesis (Welbaum et al., 1998). It is considered that seed germination vigor and longevity are correlated to molecular mobility, M.E. Hudson, S.H. Chang, B. Han, X. Wang, and T. Zhu, Torrey Mesa Research Institute, Syngenta Research and Technology, 3115 Merryfi eld Row, San Diego, CA 92121. M.E. Hudson, Dep. of Crop Sciences, Univ. of Illinois, 334 NSRC, 1101 W. Peabody Blvd., Urbana, IL 61801. W. Yu and T. Zhu, Syngenta Biotechnology, 3054 Cornwallis Rd., Research Triangle Park, NC 27709. T. Bruggink and P. van der Toorn, Syngenta Seeds B.V., Westeinde 62, Enkhuizen, the Netherlands. Received 1 Dec. 2006. *Corresponding author ([email protected]). Published in Crop Sci. 47(S2) S96–S112. Published 14 July 2007. doi:10.2135/cropsci2006.12.0758tpg © Crop Science Society of America 677 S. Segoe Rd., Madison, WI 53711 USA Abbreviations: EDTA, ethylenediaminetetraacetic acid; FDR, false discovery rate; MES, 2-N-morpholino-ethane-sulfonic acid; MOID, match-only integral distribution; RH, relative humidity; PCR, polymerase chain reaction; RT–PCR, reverse transcription polymerase chain reaction; SAM, Statistical Analysis of Microarrays; SAPE, Streptavidin Phycoerythrin. Ge ome The Original Research Published online July 16, 2007