Capillary electrophoresis of PCR fragments with 5´-labelled primers for testing the SARS-Cov-2.
J Virol Methods
; 284: 113937, 2020 10.
Article
in English
| MEDLINE | ID: covidwho-638504
Preprint
This scientific journal article is probably based on a previously available preprint. It has been identified through a machine matching algorithm, human confirmation is still pending.
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This scientific journal article is probably based on a previously available preprint. It has been identified through a machine matching algorithm, human confirmation is still pending.
See preprint
ABSTRACT
Due to the huge demand for SARS-Cov-2 determination,alternatives to the standard qtPCRtestsare potentially useful for increasing the number of samples screened. Our aim was to develop a direct fluorescent PCR capillary-electrophoresis detection of the viral genome. We validated this approach on several SARS-Cov-2 positive and negative samples.We isolated the naso-pharingealRNA from 20 positive and 10 negative samples. The cDNA was synthesised and two fragments of the SARS-Cov-2 were amplified. One of the primers for each pair was 5´-end fluorochrome labelled. The amplifications were subjected to capillary electrophoresis in ABI3130 sequencers to visualize the fluorescent peaks.The two SARS-Cov-2 fragments were successfully amplified in the positive samples, while the negative samples did not render fluorescent peaks. In conclusion, we describe and alternative method to identify the SARS-Cov-2 genome that could be scaled to the analysis of approximately 100 samples in less than 5 h. By combining a standard PCR with capillary electrophoresis our approach would overcome the limits imposed to many labs by the qtPCR and increase the testing capacity.
Keywords
Full text:
Available
Collection:
International databases
Database:
MEDLINE
Main subject:
Pneumonia, Viral
/
Polymerase Chain Reaction
/
Coronavirus Infections
/
Electrophoresis, Capillary
/
Clinical Laboratory Techniques
/
Betacoronavirus
Type of study:
Diagnostic study
/
Prognostic study
Limits:
Humans
Language:
English
Journal:
J Virol Methods
Year:
2020
Document Type:
Article
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