关键词: RNA芯片杂交, 斑点杂交, cDNA芯片, 马铃薯纺锤块基类病毒(PSTVd)

Abstract:

Potato diseases caused by potato spindle tuber viroid(PSTVd) bring about substantial losses in agricultural production. So developing a high throughput, rapid, sensitive and specific detection method is necessary. This article reports a new method to detect plant viroid——RNA glass slide hybridization. This method combines dot blot hybridization technique with cDNA microarray technique. We could spot a number of RNA samples extracted from new leaves of potatoes onto a silanated glass slide directly and then perform the hybridization on the slide with Cy5-labeled probe of PSTVd prepared by PCR. Analyzing the results of hybridization, we could ascertain which sample is infected by PSTVd. The factors affecting the results were optimized and used to test six potato samples with PSTVd-specific probes. The developed method was used to detect two samples which infected by PSTVd. These results were in good accordance with those from the RT-PCR approach and the samples are proved to be PSTVd by cloning and sequencing. So it is an accurate and reliable method for detecting plant viroid and the first report for the diagnosis of PSTVd using dot blot hybridization and RNA glass side hybridization with fluorescently labeled probe.

Key words: RNA glass slide hybridization, Dot blot hybrid ization, cDNA microarray, Potato spindle tuber viroid(PSTVd)