Hs. Chu et Tj. Cho, ISOLATION OF SALICYLIC ACID-INDUCED GENES IRE BRASSICA-NAPUS BY SUBTRACTIVE HYBRIDIZATION, Molecules and cells, 6(6), 1996, pp. 766-772
Subtractive hybridization has been performed to isolate salicylic acid
-induced genes in Brassica napus, as a first step toward understanding
plant defense mechanisms. In this procedure, tester cDNA was adapted
with a linker and hybridized with a 10-fold excess of driver cDNA havi
ng a different linker. After hybridization, the DNA mixture was treate
d with T4 DNA polymerase and S1 nuclease so that only double-stranded
tester/tester DNAs were amplified in subsequent polymerase chain react
ions (PCR). After repeated rounds of subtractive hybridization, the en
riched DNAs were analyzed by sequencing and database search. Of the en
riched sequences examined, several putative defense-related genes have
been identified. These include cytochrome P450, glutathione S-transfe
rase, pathogenesis-related la protein, and a lipase homolog genes. Rev
erse transcription-PCR showed that these genes are induced by salicyli
c acid and/or wounding.