Detection of sars-cov-2 variant

A method for detecting a SARS-COV-2 variant, the method comprising:

• (a) amplifying a nucleic acid in a sample suspected of comprising the SARS-COV-2 variant, the variant comprising at least one insertion, deletion or single nucleotide polymorphism (SNP) as compared to a wildtype SARS-COV-2 nucleic acid, and (b) detecting an amplicon produced in (a);
• wherein amplifying the nucleic acid comprises:
• (i) performing a polymerase chain reaction using at least one set of nucleic acids (primer set), wherein the primer set amplifies a nucleic acid of the SARS-COV-2 variant comprising the insertion or deletion and the amplicon detected in (b) is larger or smaller than a corresponding amplicon generated from a wildtype SARS-COV-2 nucleic acid;
• and/or
• (ii) performing a polymerase chain reaction using a first nucleic acid primer, a second nucleic acid primer and third nucleic acid primer, wherein the first nucleic acid primer preferentially anneals to a nucleic acid comprising a mutant allele of the SNP, the second nucleic acid primer preferentially anneals to a nucleic acid comprising wildtype allele of the SNP, and the third nucleic acid allows for the amplification of a nucleic acid comprising the SNP.

A method of generating a primer for detecting a single nucleotide polymorphism (SNP) in a nucleic acid, the method comprising:

• (a) providing a first nucleic acid sequence complementary to a nucleic acid comprising a mutant allele of the SNP, wherein the first nucleic acid sequence comprises a terminal 3′ nucleotide complementary to either the mutant or the wildtype nucleotide at the location of the SNP;
• (b) substituting an adenine (A) for a thymine (T) (T>A substitution) one base pair upstream (5′ direction) of the terminal 3′ nucleotide complementary to the mutant or wildtype nucleotide at the location of the SNP to generate a mutant primer sequence or a wildtype primer sequence;
• and
• (c) generating a mutant primer comprising the mutant primer sequence and/or a wildtype primer comprising the wildtype primer sequence;
• wherein binding of the mutant primer to a wildtype allele or the binding of the wildtype primer to a mutant allele is destabilized as compared to a mutant primer or wildtype primer without the T>A substitution.

A method of generating a primer for detecting a single nucleotide polymorphism (SNP) in a nucleic acid, the method comprising:

• (a) providing a first nucleic acid sequence complementary to a nucleic acid comprising a mutant allele of the SNP, wherein the first nucleic acid sequence comprises a terminal 3′ nucleotide complementary to the mutant nucleotide at the location of the SNP;
• (b) generating a mutant primer sequence by substituting an adenine (A) for a cytosine (C) (C>A substitution) two base pairs upstream (5′ direction) of the terminal 3′ nucleotide that is complementary to the mutant allele;
• and
• (c) preparing an isolated nucleic acid primer (mutant primer) comprising the mutant primer sequence, wherein the mutant primer binds the mutant allele with a higher specificity than an equivalent primer without the C>A substitution.