A quantitative PCR, real-time RT-PCR, and one-step real-time RT-PCR using SYBR Green-based tools for reliable detection and relative quantitation of Prune dwarf virus (PDV) in stone fruits are described. The assay reliability was tested on 55 samples from different hosts and regions. The sensitivity of the assay was also compared with other assays with different primers. Two plant-expressed genes, actin and 18S rRNA, were used as housekeeping genes for accurate quantitation of PDV in stone fruit trees. The expression of the gene for actin and the 18S ribosomal RNA gene corresponded with each other accurately, with standard deviation values of 1.905 cycles on average, 1.36 for Prunus persica, and 2.45 for other Prunus species tested. The results of this study support the need to use more than one housekeeping gene as an internal control to avoid possible errors caused by unstable internal control gene mRNA expression when quantifying the extent of PDV infection.
- MeSH
- Actins genetics MeSH
- Staining and Labeling methods MeSH
- Fluorescent Dyes MeSH
- Ilarvirus genetics isolation & purification MeSH
- Organic Chemicals MeSH
- Reverse Transcriptase Polymerase Chain Reaction methods standards MeSH
- Reference Standards MeSH
- RNA, Ribosomal, 18S genetics MeSH
- RNA, Plant genetics MeSH
- RNA, Viral genetics MeSH
- Plant Proteins genetics MeSH
- Prunus virology MeSH
- Viral Load MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
