Method of detecting or quantitating endogenous wheat dna and method of determining contamination rate of genetically modified wheat in test sample

Abstract

An object of the present invention is to discover an endogenous wheat sequence satisfying the conditions of: a) it is universally present in varieties of wheat, b) the amount present (detected amount) is not affected depending on the wheat variety, c) even if other grains are present, only wheat can be detected without cross-reactivity, and d) it is amplified quantitatively by the PCR reaction. A further object of the present invention is to provide a method of accurately detecting and quantitating endogenous wheat DNA in a test sample by the polymerase chain reaction. The present invention provides replicable DNA comprising: a DNA sequence as endogenous DNA common to both genetically modified wheat and non-genetically modified wheat; and one or more DNA sequence each comprising a sequence that is specific to a strain of genetically modified wheat.

1 .- 13 . (canceled) 
     
     
         14 . Replicable DNA comprising:
 a DNA sequence consisting of the nucleotide sequence identified as SEQ ID NO: 2 or a partial sequence thereof as endogenous DNA common to both genetically modified wheat and non-genetically modified wheat; and   one or more DNA sequences each comprising a sequence that is specific to a strain of genetically modified wheat.   
     
     
         15 . Replicable DNA comprising:
 a DNA sequence consisting of a nucleotide sequence having at least 80% homology with SEQ ID NO: 2 or a partial sequence thereof as endogenous DNA common to both genetically modified wheat and non-genetically modified wheat; and   one or more DNA sequences each consisting of a sequence that is specific to a strain of genetically modified wheat.   
     
     
         16 . The DNA according to  claim 14 , wherein the DNA sequence consisting of a nucleotide sequence having at least 80% homology with SEQ ID NO: 2 or a partial sequence thereof is selected from SEQ ID NOs: 11 to 18. 
     
     
         17 . Replicable DNA comprising:
 a DNA sequence consisting of a nucleotide sequence capable of being amplified by the polymerase chain reaction using a primer pair chosen from primer pairs (i) to (ix), below, as endogenous DNA common to both genetically modified wheat and non-genetically modified wheat; and   one or more DNA sequences each consisting of a nucleotide sequence that is specific to a strain of genetically modified wheat,   wherein the primer pairs (i) to (ix) are:   (i) a primer pair consisting of a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 3 and a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 4;   (ii) a primer pair consisting of a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 5 and a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 4;   (iii) a primer pair consisting of a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 3 and a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 6;   (iv) a primer pair consisting of a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 5 and a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 6;   (v) a primer pair consisting of a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 3 and a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 7;   (vi) a primer pair consisting of a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 3 and a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 8;   (vii) a primer pair consisting of a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 5 and a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 8;   (viii) a primer pair consisting of a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 5 and a nucleic acid molecule comprising the nucleotide sequence identified as SEQ ID NO: 7; and   (ix) a primer pair consisting of a pair of nucleic acid molecules, wherein one nucleic acid molecule in the primer pair in (ix) comprises a nucleotide sequence in common with at least 80% continuous nucleotide sequence of one nucleic acid molecule in one of the primer pairs in (i) to (viii) above, and the other nucleic acid molecule in the primer pair in (ix) comprises a nucleotide sequence in common with at least 80% continuous nucleotide sequence of the other nucleic acid molecule in the same primer pair in (i) to (viii) above.   
     
     
         18 . The DNA according to  claim 17 , wherein the DNA sequence consisting of a nucleotide sequence capable of being amplified by the polymerase chain reaction is chosen from SEQ ID NOs: 11 to 18.