Download Gene Expression Patterns for the Malting Barley `Alexis`

Gene Expression Patterns for the Malting Barley ‘Alexis’
Loraine Watson and Robert J. Henry
CRCMPB Centre for Plant Conservation Genetics Southern Cross University Lismore NSW 2480
Over the past three years we have undertaken a novel project using Microarray analysis. The impetus behind our research
has been the importance of germination rate in the production of high quality malt. The aim of our study was to examine the
genetic mechanisms of germination in malting barley varieties. Here, we present a subset of data - outlining gene expression
pattern results that may be of particular importance for understanding the genetic triggers involved in barley germination.
A cDNA library was constructed using equal quantities of RNA collected from embryos of the malting barley variety ‘Alexis’. Embryos were dissected away from the starchy endosperm
and frozen in liquid nitrogen prior to RNA extraction. The stages of development chosen for inclusion were embryos that had been incubated for 24 to 96hr since imbibition. This time
period was considered to most closely represent the malting process.
Germinating ‘Alexis’ barley at a) 24hr; b) 48hr;
c) 72hr; and d) 96hr post-imbibition.
Results
‘Alexis’ cDNA Microarrays were constructed using equal quantities of RNA taken from 24hr, 48hr, 72hr and 96hr postimbibition time points. Each slide contains two copies of the 4945-spot array. The Microarrays also contain ‘Lucidea Universal
Scorecard’® controls from Amersham Biosciences to enable inter-slide comparisons. The arrays were hybridised with RNA
collected from a number of barley varieties at different stages of germination to test for differences in gene expression patterns.
Microarray hybridisation data has been processed using ImaGeneTM software and is currently undergoing statistical analysis using GeneSpring v. 6.0 (Silicon Genetics).
1. ‘Alexis’ time course experiment gene
expression profile graph of embryos collected
at 8hr, 48hr, 72hr, and 96hr post-imbibition,
compared to the gene expression of embryos
collected at 24hr post-imbibition.
4. Subset of ‘Galleon’ vs. ‘Alexis’ time course showing genes that
appear to be down-regulated in ‘Galleon’ with respect to ‘Alexis’ at
48hr post-imbibition. ‘Galleon’ was observed to have a slower
germination rate than ‘Alexis’ under laboratory conditions.
Future Directions
Further statistical analysis of the Microarray data will provide firm
genetic candidates with relevance to germination rate. These
candidates will be sequenced and subjected to Real Time PCR to
both verify the Microarray data and quantify the level of
expression.
Acknowledgements
3. Subset of ‘Alexis’ time course showing
genes with lower expression at 8hr postimbibition and much higher expression by
48hr post-imbibition.
The project is funded by the CRCMPB who are gratefully acknowledged.
We would also like to thank staff at the Plant Biotechnology Centre, La
Trobe University, Victoria, for their assistance with Microarray
construction and hybridisation. The advice of Dr Timothy Holton on
cDNA library construction is also acknowledged with gratitude.
Produced by SCU, Marketing & External Relations Directorate. 1307
2. Subset of ‘Alexis’ time course showing
genes that appear to be up-regulated at 8hr
post-imbibition, down-regulated at 48hr postimbibition and then up-regulated once more
by 72hr post-imbibition.