Dataset Information

Detection of the Janus kinase 2 V617F mutation using a locked nucleic-acid, real-time polymerase chain reaction assay.

ABSTRACT: The purpose of this study was to develop a real time polymerase chain reaction (PCR) assay for the detection of the JAK2 V617F mutation that could be used in diagnostic laboratories. Sanger sequencing and a newly developed locked nucleic-acid, real-time PCR assay were used to detect the JAK2 V617F mutation. There was 100% agreement between the sequencing and PCR analysis. Both assays were able to detect the mutation in all 24 of the 60 test specimens harbouring the mutation.

SUBMITTER: Senamela T

PROVIDER: S-EPMC5806058 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Detection of the Janus kinase 2 V617F mutation using a locked nucleic-acid, real-time polymerase chain reaction assay.

Senamela Tshiphiri T Kock Marleen M Becker Piet P Potgieter Joachim J C JJC

African journal of laboratory medicine 20180131 1

The purpose of this study was to develop a real time polymerase chain reaction (PCR) assay for the detection of the <i>JAK2</i> V617F mutation that could be used in diagnostic laboratories. Sanger sequencing and a newly developed locked nucleic-acid, real-time PCR assay were used to detect the <i>JAK2</i> V617F mutation. There was 100% agreement between the sequencing and PCR analysis. Both assays were able to detect the mutation in all 24 of the 60 test specimens harbouring the mutation. ...[more]

PMID: 29435426

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Project description:The definitive diagnosis of lyssavirus infection (including rabies) in animals and humans is based on laboratory confirmation. The reference techniques for post-mortem rabies diagnosis are still based on direct immunofluorescence and virus isolation, but molecular techniques, such as polymerase chain reaction (PCR) based methods, are increasingly being used and now constitute the principal tools for diagnosing rabies in humans and for epidemiological analyses. However, it remains a key challenge to obtain relevant specificity and sensitivity with these techniques while ensuring that the genetic diversity of lyssaviruses does not compromise detection. We developed a dual combined real-time reverse transcription polymerase chain reaction (combo RT-qPCR) method for pan-lyssavirus detection. This method is based on two complementary technologies: a probe-based (TaqMan) RT-qPCR for detecting the RABV species (pan-RABV RT-qPCR) and a second reaction using an intercalating dye (SYBR Green) to detect other lyssavirus species (pan-lyssa RT-qPCR). The performance parameters of this combined assay were evaluated with a large panel of primary animal samples covering almost all the genetic variability encountered at the viral species level, and they extended to almost all lyssavirus species characterized to date. This method was also evaluated for the diagnosis of human rabies on 211 biological samples (positive n = 76 and negative n = 135) including saliva, skin and brain biopsies. It detected all 41 human cases of rabies tested and confirmed the sensitivity and the interest of skin biopsy (91.5%) and saliva (54%) samples for intra-vitam diagnosis of human rabies. Finally, this method was successfully implemented in two rabies reference laboratories in enzootic countries (Cambodia and Morocco). This combined RT-qPCR method constitutes a relevant, useful, validated tool for the diagnosis of rabies in both humans and animals, and represents a promising tool for lyssavirus surveillance.

Dataset Information

Detection of the Janus kinase 2 V617F mutation using a locked nucleic-acid, real-time polymerase chain reaction assay.

Publications

Detection of the Janus kinase 2 V617F mutation using a locked nucleic-acid, real-time polymerase chain reaction assay.

Similar Datasets

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