Development and validation of a flax (Linum usitatissimum L.) gene expression oligo microarray.
BACKGROUND
Flax (Linum usitatissimum L.) has been cultivated for around 9,000 years and is therefore one of the oldest cultivated species. Today, flax is still grown for its oil (oil-flax or linseed cultivars) and its cellulose-rich fibres (fibre-flax cultivars) used for high-value linen garments and composite materials. Despite the wide industrial use of flax-derived products, and our actual understanding of the regulation of both wood fibre production and oil biosynthesis more information must be acquired in both domains. Recent advances in genomics are now providing opportunities to improve our fundamental knowledge of these complex processes. In this paper we report the development and validation of a high-density oligo microarray platform dedicated to gene expression analyses in flax.


RESULTS
Nine different RNA samples obtained from flax inner- and outer-stems, seeds, leaves and roots were used to generate a collection of 1,066,481 ESTs by massive parallel pyrosequencing. Sequences were assembled into 59,626 unigenes and 48,021 sequences were selected for oligo design and high-density microarray (Nimblegen 385K) fabrication with eight, non-overlapping 25-mers oligos per unigene. 18 independent experiments were used to evaluate the hybridization quality, precision, specificity and accuracy and all results confirmed the high technical quality of our microarray platform. Cross-validation of microarray data was carried out using quantitative qRT-PCR. Nine target genes were selected on the basis of microarray results and reflected the whole range of fold change (both up-regulated and down-regulated genes in different samples). A statistically significant positive correlation was obtained comparing expression levels for each target gene across all biological replicates both in qRT-PCR and microarray results. Further experiments illustrated the capacity of our arrays to detect differential gene expression in a variety of flax tissues as well as between two contrasted flax varieties.


CONCLUSIONS
All results suggest that our high-density flax oligo-microarray platform can be used as a very sensitive tool for analyzing gene expression in a large variety of tissues as well as in different cultivars. Moreover, this highly reliable platform can also be used for the quantification of mRNA transcriptional profiling in different flax tissues.
Fenart
Stéphane
S
Université Lille Nord de France, Lille 1 UMR INRA 1281, SADV, F- 59650 Villeneuve d'Ascq cedex, France.
Ndong
Yves-Placide Assoumou
YP
Duarte
Jorge
J
Rivière
Nathalie
N
Wilmer
Jeroen
J
van Wuytswinkel
Olivier
O
Lucau
Anca
A
Cariou
Emmanuelle
E
Neutelings
Godfrey
G
Gutierrez
Laurent
L
Chabbert
Brigitte
B
Guillot
Xavier
X
Tavernier
Reynald
R
Hawkins
Simon
S
Thomasset
Brigitte
B
D020451:Contig Mapping; D019597:Flax / Q000235:genetics* / Q000254:growth & development; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant / Q000235:genetics; D005838:Genotype; D058977:Molecular Sequence Annotation; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D009928:Organ Specificity / Q000235:genetics; D018547:Plant Stems / Q000235:genetics; D025341:Principal Component Analysis; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA