Dot-blot hybridization was applied in this paper to detect five viruses infecting cucurbitaceous crops, Zuccini yellow mosaic virus （ZYMV）, Watermelon mosaic virus （WMV）, Cucumber mosaic virus （CMV）, Papaya ring...Dot-blot hybridization was applied in this paper to detect five viruses infecting cucurbitaceous crops, Zuccini yellow mosaic virus （ZYMV）, Watermelon mosaic virus （WMV）, Cucumber mosaic virus （CMV）, Papaya ringspot viruswatermelon strain （PRSV-W） and Squash mosaic virus （SqMV）, as a good alternative assay in seed health test and epidemiological and transgenic research. Digoxigenin-labelled cDNA probes of the five viruses were synthesized by PCR with the specific primers and applied in dot-blot hybridization to detect five viruses in crude extraction of the infected leaves. And three SqMV probes of different lengths （0.55, 1.6, and 2.7 kb, respectively） were designed to investigate the effect of hybridization. The results showed that the sensitivity for detecting the crude extraction of infected leaves by ZYMV, WMV, CMV, PRSV-W, and SqMV was down to 1：160, 1：160, 1：320, 1：160, and 1：320, respectively. Three SqMV probes of different length showed no differences on the sensitivity and specificity. The digoxigenin-labelled probes prepared by PCR could be used for accurate and rapid identification of 5 viruses infecting cucurbitaceous crops with good stabilities, sensitivities, specificity, and reproducibilifies.展开更多
A novel real-time RT-PCR method,BMPs based real-time RT-PCR which integrated with magne-tic separation technique of bacterial magnetic particles(BMPs),was set up for detection of Squash mosaic virus(SqMV).After SqMV p...A novel real-time RT-PCR method,BMPs based real-time RT-PCR which integrated with magne-tic separation technique of bacterial magnetic particles(BMPs),was set up for detection of Squash mosaic virus(SqMV).After SqMV particles in crude sap were concentrated by BMPs,viral RNAs were released and detected by real time RT-PCR.The results indicated that BMPs based real-time RT-PCR was efficient,and the detection sensibility was equivalent to that of the Trizol based real-time RT-PCR,of which Trizol reagent was used for viral RNAs extration.Comparing to Trizol-based method,the BMPs-based method had advantages of simplicity on operation,time saving for RNA extraction and without using noxious organic chemicals.展开更多
文摘Dot-blot hybridization was applied in this paper to detect five viruses infecting cucurbitaceous crops, Zuccini yellow mosaic virus （ZYMV）, Watermelon mosaic virus （WMV）, Cucumber mosaic virus （CMV）, Papaya ringspot viruswatermelon strain （PRSV-W） and Squash mosaic virus （SqMV）, as a good alternative assay in seed health test and epidemiological and transgenic research. Digoxigenin-labelled cDNA probes of the five viruses were synthesized by PCR with the specific primers and applied in dot-blot hybridization to detect five viruses in crude extraction of the infected leaves. And three SqMV probes of different lengths （0.55, 1.6, and 2.7 kb, respectively） were designed to investigate the effect of hybridization. The results showed that the sensitivity for detecting the crude extraction of infected leaves by ZYMV, WMV, CMV, PRSV-W, and SqMV was down to 1：160, 1：160, 1：320, 1：160, and 1：320, respectively. Three SqMV probes of different length showed no differences on the sensitivity and specificity. The digoxigenin-labelled probes prepared by PCR could be used for accurate and rapid identification of 5 viruses infecting cucurbitaceous crops with good stabilities, sensitivities, specificity, and reproducibilifies.
文摘A novel real-time RT-PCR method,BMPs based real-time RT-PCR which integrated with magne-tic separation technique of bacterial magnetic particles(BMPs),was set up for detection of Squash mosaic virus(SqMV).After SqMV particles in crude sap were concentrated by BMPs,viral RNAs were released and detected by real time RT-PCR.The results indicated that BMPs based real-time RT-PCR was efficient,and the detection sensibility was equivalent to that of the Trizol based real-time RT-PCR,of which Trizol reagent was used for viral RNAs extration.Comparing to Trizol-based method,the BMPs-based method had advantages of simplicity on operation,time saving for RNA extraction and without using noxious organic chemicals.
