HLVd Detection Kits

PDD-356

HLVd Detection Kits

HLVd RT-PCR Kit is designed for the detection of Hop Latent Viroid specific RNA in a real-time PCR.

HLVd RT-PCR Kit is designed for the detection of HLVd-specific RNA in a real-time PCR based on the use of technology. This kit is designed for research use only and not for use in diagnostic procedures. The detection of HLVd-specific RNA is based on one-step RT-PCR providing a simple, reliable, and rapid result for the detection of HLVd infection. HLVd RT-PCR Kit includes a PCR control to monitor for PCR inhibition and to validate the quality of the sample and the detection result. The HLVd RT-PCR Kit comprises a Master Mix for the target and PCR control detection, Primer & Probe Mix, as well as a positive control and negative control (nuclease-free water) to confirm the integrity of the kit reagents.

1. Before use, suitable amounts of all components should be completely thawed at room temperature, mixed by gentle vortexing or by pipetting, and centrifuged briefly.
2. Work quickly on ice.
3. The recommended minimum number of DNA samples tested per PCR run is 6.

Viroid

1. Follow universal precautions. All specimens should be considered as potentially infectious and handled accordingly.
2. Ensure that a suitable lab coat, disposable gloves, and protective goggles are worn when handling specimens and kit reagents.
3. Use sterile pipette tips with filters. Use proper pipetting techniques and maintain the same pipetting pattern throughout the procedure to ensure optimal and reproducible values.
4. Ensure that appropriate specimen collection, transport, storage, and processing techniques are followed for optimal performance of this test.

1. MDx 2X RT-PCR Master Mix 350 µL
2. HLVd Primer & Probe Mix 70 µL
3. HLVd Positive Control 50 µL
4. Nuclease-Free Water (Negative control) 1.25 mL
5. Product Insert 1

1. Appropriate Real-Time PCR Instrument with FAM and HEX filter channel
2. DNA Purification Kit
3. Disposable powder-free gloves
4. Benchtop microcentrifuge
5. Micropipettors
6. Sterile pipette tips with filters
7. PCR tubes
8. Vortex mixer
9. PCR reaction preparation station (Optional)

Hop latent viroid (HLVd) is a single-stranded, circular infectious RNA of 256 nucleotides. Initially it was believed that this viroid did not produce any symptoms of disease, thus the inclusion of the term latent in the viroid name. However it is now known that HLVd infection occurs worldwide in hop (Humulus lupulus) and is deleterious to production in certain cultivars. Quantitative changes in some secondary metabolites in the secretory glands forming lupulin are induced by HLVd infection, indicating that the viroid is not truly latent. Hop Latent Viroid infection depends on the metabolism of the host plant, and it’s replication and pathogenesis are mediated by its structural features and its ability to interact with cellular factors.

PCR

The specificity of HLVd RT-PCR Kit is first and foremost ensured by the selection of the HLVd-specific primers, as well as the selection of stringent reaction conditions. The HLVd primers were checked for possible homologies to all plant viroids in GenBank published sequences by sequence comparison analysis.

All kit components should be stored at -20°C upon arrival. Repeated thawing and freezing (> 2 x) of the Master Mix and Positive Control should be avoided, as this may affect the performance of the assay. If the reagents are to be used only intermittently, they should be frozen in aliquots. All reagents can be stored for 1 year at -20°C without showing any reduction in performance.