Functional Genomics of Developing Barley Seeds
Expressed Sequence Tags (ESTs), typically single-pass and partial sequences from cDNA clones, directly represent the transcriptome or transcribed portions of the genome and avoid the highly repetitive DNA that makes up the bulk of genomic DNA. Within the EST set, sequences generated from cDNA libraries of developing seeds are represented with 47,066 ESTs. Along with other EST collections available at IPK from different tissues at different developmental stages in silico expression expression analysis were peformed. This digital in silico method has first been implemented to assess genes preferentially expressed in seeds (http://gene-exp.ipk-gatersleben.de/c5.html).
The major goals of the project are the provision of the necessary material (mainly cDNAs and high density arrays thereof) and the establishment of the technologies to allow a systematic analysis of gene expression during barley seed development.
cDNA libraries were produced from the early caryopsis development 0-12 DAF (http://pgrc.ipk-gatersleben.de). In the first phase we prepared cDNA macroarray filter with 700 clones representing mostly from early caryopsis development (part of unigene set from HY cDNA library). In the second phase, almost 1,440 clones representing mostly unique sequences from early caryopsis development were selected to create a high density cDNA macroarray filter (unigene set from HY cDNA library). The macroarrays were hybridised to 33P-labeled cDNA probes derived from dissected maternal and filial (contains endopserm and embryo) tissue in two day intervals (0to 12 DAF). Spatial and temporal mRNA profiles characterizing the developing maternal and filial part of the barley grain during pre-storage and the early storage phase were depicted. In co-operation, 1,400 array has been used to study seed germination and malting-related traits in barley.
In the third generation, 12,000 unigene set has been identified based on the developing seed barley EST dataset. As a result, a set comprising 12K unique sequences was used for preparation of high-density cDNA array. This 12K array provides a valuable resource for studying complete “Brenda” seed development.
Contact: Dr. Nese Sreenivasulu
Scientific Investigators: N. Sreenivasulu, V. Radchuk, W. Weschke, M. Strickert
Project Co-Ordinator: Prof. Ulrich Wobus
Data Management for the WWW and Web Responsibility: U. Scholz