The ligation amplification reaction (LAR)--amplification of specific DNA sequences using sequential rounds of template-dependent ligation.
A novel DNA sequence detection method that utilizes the ligation of oligonucleotide pairs that are complementary to adjacent sites on appropriate DNA templates is described. The product is increased by either linear or exponential amplification using sequential rounds of template-dependent ligation. In the case of linear amplification, a single pair of oligonucleotides is ligated, the reaction is heated to dissociate the ligation product, and an additional round of ligation is performed. After n rounds there is a (1 + x) X n-fold amplification of product, where x is the efficiency of the ligation reaction. Exponential amplification utilizes two pairs of oligonucleotides, one complementary to the upper strand and one to the lower strand of a target sequence. The products of the ligation reaction serve as templates for subsequent rounds of ligation. In this case there is (1 + x)(n-1)-fold amplification of product after n rounds. A single base-pair mismatch between the annealed oligonucleotides and the template prevents ligation, thus allowing the distinction of single base-pair differences between DNA templates. At high template concentrations, the ligation reaction has an efficiency approaching 100%. In this report, we demonstrate the use of the ligation amplification reaction (LAR) to distinguish the normal from the sickle cell allele of the human beta-globin gene. We also report the use of LAR as a detection system for polymerase chain reaction-enriched DNA sequences.
Wu
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Department of Molecular Biochemistry, Beckman Research Institute of the City of Hope, Duarte, California 91010.
Wallace
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RB
D000755:Anemia, Sickle Cell / Q000235:genetics; D001483:Base Sequence; D004247:DNA / Q000032:analysis* / Q000235:genetics; D011088:DNA Ligases; D004252:DNA Mutational Analysis; D005784:Gene Amplification; D005914:Globins / Q000235:genetics; D006451:Hemoglobin, Sickle / Q000235:genetics; D006801:Humans; D015345:Oligonucleotide Probes; D013698:Templates, Genetic
D006451:Hemoglobin, Sickle; D015345:Oligonucleotide Probes; D005914:Globins; D004247:DNA; D011088:DNA Ligases