SYSTEM AND METHOD FOR DETERMINING KNOWN DNA VARIANTS WITH TEMPERATURE GRADIENT ELECTROPHORESIS

摘要

A first aspect of the invention relates to a method of determining the genotype of a sample polynucleotide having at least a first variant site. The method may include amplifying at least a portion of the sample polynucleotide to obtain first amplicons, the first amplicons including the first variant site. The position of the first variant site along the sample nucleotide is preferably known. The first amplicons may be combined with first and second different polynucleotide controls. The first and second polynucleotide controls may differ by at least one base therealong, the position of the at least one differing base preferably corresponds to the position of the first variant site along the sample polynucleotide. A plurality of first duplexes may be prepared. At least some and preferably each of at least some of the first duplexes may include (i) a polynucleotide strand of one of the first amplicons and (ii) a complementary polynucleotide strand of the first polynucleotide control. A plurality of second duplexes may be prepared. At least some and preferably each of at least some of the second duplexes including (i) a polynucleotide strand of one of the first amplicons and (ii) a complementary polynucleotide strand of the second polynucleotide control. The first and second duplexes may be subjected to temperature gradient electrophoresis (TGE) to obtain first and second electrophoresis data, which are indicative of the genotype of the first variant site. The genotype of the first variant site of the sample polynucleotide may be determined based on the first and second electrophoresis data. The determination of the genotype may include determining a number of peaks present in the first electrophoresis data and a number of peaks present in the second electrophoresis data.