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Title: SURVEY OF BTH-INDUCED GENES IN PAPAYA BY SUPPRESSION SUBTRACTIVE HYBRIDIZATION

Author
item Albert, Henrik
item QIU, XIAOHUI - U.H. MANOA
item WANG, MINGLI - HARC
item Moore, Paul

Submitted to: Plant Genomics 2003
Publication Type: Abstract Only
Publication Acceptance Date: 8/15/2003
Publication Date: 10/10/2003
Citation: Albert, H.H., Qiu, X., Wang, M., Moore, P.H. 2003. Survey of BTH-induced genes in papaya by suppression subtractive hybridization. West Sect ASPB Oct. 10-11. Plant Genomics 2003: p. 21. 2003.

Interpretive Summary:

Technical Abstract: Papaya has been shown to respond to pathogen challenge and benzothiadiazole(BTH) treatment with an increase in B-glucanase and chitinase enzyme activities[1]. We now report a survey of BTH-induced papaya genes using suppression subtractive hybridization(SSH)[2,3]. Twenty-five unique ESTs representing 23 genes were confirmed by reverse-northern and quantitative (phosphor-imager) northern blots to be induced (greater than or equal to 1.5 fold), obtained from 332 tested SSH clones. mRNA accumulation for these ESTs was further analyzed by qRT-PCR, and 18 were confirmed to be induced greater than or equal to 1.5 fold. Additionally, six SSH ESTs undetectable by northern blot, four papaya PR-1 genes, and NPR1, were analyzed by qRT-PCR. PR-1a and c were found to be down-regulated by BTH, while PR-1b and PR-1d were induced; PR-1d approx. 20-fold. NPR1 was induced approx. 1.7 fold, comparable to the level reported for arabidopsis. All six ESTs undetectable by northern blot were detected by qRT-PCR and one of these was induced. BTH induced genes included numerous "expected" genes (e.g. a peroxidase, two chitinases and an osmotin-like protein) but also some unexpected genes, including homologs of citrus blight-associated P12 protein (induced approx. 6-fold) and a putative arabidopsis Fe(II)/ascorbate oxidase (induced approx. 6000-fold). Neither of the induced PR-1 genes was recovered in our SSH sample. While SSH was found to be an effective method for studying a "non-model" system, it did not identify a complete list of BTH-induced genes. For confirmation of induction, reverse-northern blots were found to be efficient and relatively reliable. For final confirmation, qRT-PCR was found to be more sensitive than northern blots. 1. Zhu, Y.J., et al. Acta Horticulturae, 2002. 576: p. 475-481 2. Diatchenko, L., et al. Methods Enzymol, 1999. 303: p. 349-80 3. Diatchenko, L., et al. Proc Natl Acad Sci USA, 1996. 93(12):p. 6025-30.