Nucleic acid quantification method and reagent for quantifying nucleic acid

A nucleic acid quantification method for quantifying a proportion of a mutant-type base sequence to a wild-type base sequence contained in a sample, comprising:

• a step of hybridizing, to a wild-type polynucleotide containing the wild-type target base sequence and a mutant-type polynucleotide containing the mutant-type target base sequence, a wild-type probe set including a plurality of wild-type probes complementary to a wild-type target base sequence and a fragment complementary to an adjacent base sequence adjacent to a 5′-terminal side of the target base sequence and a mutant-type probe set including a mutant-type probe complementary to a mutant-type target base sequence and a fragment complementary to an adjacent base sequence adjacent to a 5′-terminal side of the target base sequence;
• a step of producing a wild-type polynucleotide including the wild-type target base sequence, by ligating the wild-type probe and the fragment hybridized to the wild-type polynucleotide and producing a wild-type polynucleotide including the wild-type target base sequence by ligating the mutant-type probe and the fragment hybridized to the mutant-type polynucleotide;
• and
• a step of fractionating the wild-type polynucleotide and the mutant-type polynucleotide by electrophoresis to determine a proportion of the mutant-type polynucleotide to the wild-type polynucleotide, wherein
• the wild-type probe has a base sequence complementary to the wild-type target base sequence at a 3′-terminal side,
• the mutant-type probe has a base sequence complementary to the mutant-type target base sequence at a 3′-terminal side, and
• the plurality of wild-type probes have mutually different molecular structures that result in different mobilities in the electrophoresis.

A reagent for quantifying nucleic acid that quantifies a proportion of a mutant-type base sequence to a wild-type base sequence contained in a sample, comprising:

• a plurality of wild-type forward primers complementary to a wild-type target base sequence;
• a mutant-type forward primer complementary to a mutant-type target base sequence;
• and
• a reverse primer complementary to a complementary strand of the target base sequence, wherein
• the wild-type forward primer has a base sequence complementary to the wild-type target base sequence at a 3′-terminal side,
• the mutant-type forward primer has a base sequence complementary to the mutant target base sequence at a 3′-terminal side, and
• the plurality of wild-type forward primers have mutually different molecular structures that result in different mobilities in the electrophoresis.

A reagent for quantifying nucleic acid that quantifies a proportion of a mutant-type base sequence to a wild-type base sequence contained in a sample, comprising:

• a plurality of wild-type probes complementary to a wild-type target base sequence;
• a plurality of mutant-type probes complementary to a mutant-type target base sequence;
• and
• a fragment complementary to an adjacent base sequence adjacent to a 5′-terminal side of the target base sequence, wherein
• the wild-type probe has a base sequence complementary to the wild-type target base sequence at a 3′-terminal side,
• the mutant-type probe has a base sequence complementary to the mutant-type target base sequence at a 3′-terminal side, and
• the plurality of wild-type probes have mutually different molecular structures that result in different mobilities in the electrophoresis.