Adaptation of a filter paper method for RNA template preparation for the detection of avocado sunblotch viroid by reverse transcription qPCR. 2022

Lara-Simone Pretorius, and Kerri A Chandra, and Anna E C Jooste, and Lebogang C Motaung, and Louisamarie E Parkinson, and Andrew D W Geering
Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, St. Lucia, QL 4072, Australia.

An easy, rapid and inexpensive method of preparing RNA template for a reverse transcription qPCR assay for avocado sunblotch viroid (ASBVd) is described. This method depends on the principle of reversible binding of viroid RNA to filter paper under different concentrations of monovalent cation. Lysis buffers containing either sodium chloride or lithium chloride were compared, and 1.5 M lithium chloride was shown to be optimal for the adsorption of the viroid RNA to the filter paper. The extraction method was validated using field samples and equivalent yields of viroid RNA were obtained using this method and either a commercial RNA extraction kit or a dsRNA chromatography method. The filter paper method of RNA extraction is ideally suited for the large-scale surveillance for ASBVd.

UI MeSH Term Description Entries
D010942 Plant Viruses Viruses parasitic on plants. Phytophagineae,Plant Virus,Virus, Plant,Viruses, Plant
D012367 RNA, Viral Ribonucleic acid that makes up the genetic material of viruses. Viral RNA
D014772 Viroids A group of pathogens comprising the smallest known agents of infectious disease. They are unencapsulated and are capable of replicating autonomously in susceptible cells. Positively identified viroids composed of single-stranded RNA have been isolated from higher plants, but the existence of DNA viroids pathogenic to animals is suspected. Viroid
D048348 Reverse Transcription The biosynthesis of DNA carried out on a template of RNA. Transcription, Reverse
D027421 Persea A plant genus in the LAURACEAE family. The tree, Persea americana Mill., is known for the Avocado fruit, the food of commerce. Avocado,Persea americana,Avocados,Persea americanas,Perseas,americana, Persea

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