DIFFERENTIATION OF THE SPOILAGE YEAST ZYGOSACCHAROMYCES-BAILII FROM OTHER ZYGOSACCHAROMYCES SPECIES USING 18S RDNA AS TARGET FOR A NONRADIOACTIVE LIGASE DETECTION REACTION
S. Stubbs et al., DIFFERENTIATION OF THE SPOILAGE YEAST ZYGOSACCHAROMYCES-BAILII FROM OTHER ZYGOSACCHAROMYCES SPECIES USING 18S RDNA AS TARGET FOR A NONRADIOACTIVE LIGASE DETECTION REACTION, Letters in applied microbiology, 19(4), 1994, pp. 268-272
A non-radioactive PCR coupled ligase detection reaction was developed
to discriminate the food spoilage yeasts Zygosaccharomyces bailii and
Z. bisporus from each other and from other members of the genus. A sho
rt region of the 18S rRNA gene was amplified from boiled cell lysates
and polymerase chain reaction (PCR) products used as target in the tem
plate directed ligation of two adjacent oligonucleotides. Ligated prod
ucts were captured using biotin-streptavidin chemistry and detected us
ing digoxigenin immuno-chemiluminescence. The ligase detection reactio
n was able to discriminate to the species level, targeting a single ba
se deletion. The specificity of the reaction was assessed using seven
species of the genus Zygosaccharomyces. Only strains of Z. bailii and
Z. bisporus gave positive results with their respective primer sets. T
he lower detection limit of the strategy was 10pg (3 x 10(7) targets)
of amplified product.