Heated oligonucleotide ligation assay (HOLA): an affordable single nucleotide polymorphism assay.
Most single nucleotide polymorphism (SNP) detection requires expensive equipment and reagents. The oligonucleotide ligation assay (OLA) is an inexpensive SNP assay that detects ligation between a biotinylated "allele-specific detector" and a 3' fluorescein-labeled "reporter" oligonucleotide. No ligation occurs unless the 3' detector nucleotide is complementary to the SNP nucleotide. The original OLA used chemical denaturation and neutralization. Heated OLA (HOLA) instead uses a thermal stable ligase and cycles of denaturing and hybridization for ligation and SNP detection. The cost per genotype is approximately US$1.25 with two-allele SNPs or approximately US$1.75 with three-allele SNPs. We illustrate the development of HOLA for SNP detection in the Early Trypsin and Abundant Trypsin loci in the mosquito Aedes aegypti (L.) and at the a-glycerophosphate dehydrogenase locus in the mosquito Anopheles gambiae s.s.
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Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523-1682, USA.
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D000330:Aedes / Q000235:genetics*; D000818:Animals; D000852:Anopheles / Q000235:genetics*; D001483:Base Sequence; D003365:Costs and Cost Analysis / Q000191:economics; D005787:Gene Frequency / Q000235:genetics; D017344:Genes, Insect / Q000235:genetics; D005828:Genetics, Population / Q000191:economics / Q000379:methods*; D005838:Genotype; D005993:Glycerolphosphate Dehydrogenase / Q000235:genetics; D006358:Hot Temperature; D008302:Mali; D008800:Mexico; D008967:Molecular Biology / Q000191:economics / Q000379:methods*; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000737:chemistry / Q000378:metabolism; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D018807:Polymorphism, Single-Stranded Conformational
D009841:Oligonucleotides; D005993:Glycerolphosphate Dehydrogenase