Improved SARS-CoV-2 PCR detection and genotyping with double-bubble primers.
Biotechniques
; 71(6): 587-597, 2021 12.
Article
in English
| MEDLINE | ID: covidwho-1406765
ABSTRACT
A new approach for improved RT-PCR is described. It is based on primers designed to form controlled stem-loop and homodimer configurations, hence the name 'double-bubble' primers. The primers contain three main regions for efficient RT-PCR a 3' short overhang to allow reverse transcription, a stem region for hot start and a template-specific region for PCR amplification. As proof of principle, GAPDH, SARS-CoV-2 synthetic RNA and SARS-CoV-2 virus-positive nasopharyngeal swabs were used as templates. Additionally, these primers were used to positively confirm the N501Y mutation from nasopharyngeal swabs. Evidence is presented that the double-bubble primers offer fast, specific, robust and cost-effective improvement in RT-PCR amplification for detection of gene expression in general and for diagnostic detection and genotyping of SARS-CoV-2 in particular.
Keywords
Full text:
Available
Collection:
International databases
Database:
MEDLINE
Main subject:
Polymerase Chain Reaction
/
COVID-19 Nucleic Acid Testing
/
SARS-CoV-2
/
COVID-19
Type of study:
Diagnostic study
/
Experimental Studies
Limits:
Humans
Language:
English
Journal:
Biotechniques
Year:
2021
Document Type:
Article
Affiliation country:
Btn-2021-0063
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