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ISSN
1672-9145
Acta Biochim Biophys Sin
2004, 36(10): 695–700
CN 31-1940/Q
Construction of the Seed-Coat cDNA Microarray and Screening of
Differentially Expressed Genes in Barley
Jin-Song PANG*, Meng-Yuan
HE, and Bao LIU
Laboratory of Molecular Epigenetics, Institute of Genetics and
Cytology, Northeast Normal University, Changchun 130024, China
Abstract Some barley mutants can
synthesize neither anthocyanins nor proanthocyanidins in the seed coat, which
is related to several genes in locus Ant13, but the exact model of action remains
unknown. We used the cDNA microarray technology with barley transcription-deficient
mutant (ant13-152) that does not synthesize proanthocyanidins as the
tester, and its wild type genotype (Triumph) as the driver, to study this
question. Six-thousand and forty-eight clones from the wild type Morex
testa+pericarp cDNA library were amplified using PCR, and the DNA fragments
were spotted on commercial amino-modified glass slide as microarray. The mRNAs
from the developing seed coat (8–15 days) of both the mutant and the wild-type
barley plants were isolated, and labeled respectively with Cy3-dUTP and
Cy5-dUTP when reversely trans-cribed to cDNAs. The labeled cDNAs were used as
probes, mixed at the same molar concentration, and hybridized with the DNA
fragments on the slide. Seventy clones exhibiting marked differential expression
(ratio>4) were identified from the microarray. All the 25 cDNA clones that
showed an over-expression in wild type in comparison to the mutant ant13-152
were sequenced. It was found that most of these over-expressing clones were
transcription/translation and hordein-associated genes. These results have laid
a solid material basis for further elucidation of the metabolic pathway in
proanthocyanidin synthesis in barley and likely other plants.
Key words barley; seed coat;
proanthocyanidin; cDNA microarray
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Received: July 5, 2004 Accepted: August 16,
2004
This work was supported by a grant from the National Science Fund
for Distinguished Young Scholars of China (30225003)
*Corresponding author: Tel, 86-431-5099367; E-mail, [email protected]