Efficient genotyping of Schistosoma mansoni miracidia following whole genome amplification.
Small parasites and larval stages pose a problem for molecular analyses because limited amounts of DNA template are available. Isothermal methods for faithfully copying DNA have the potential to revolutionize studies of such organisms. We evaluated the fidelity of multiple displacement amplification (MDA) for amplifying DNA extracted from a single miracidium of Schistosoma mansoni. To do this we genotyped DNA extracted from 28 F1 miracidia following MDA using 56 microsatellite markers. Because these miracidia were obtained from a cross between a male and female worm of known genotypes, we were able to predict the alleles present in the progeny and quantify the genotyping error rate. We found just 8/1568 genotypes deviated from Mendelian expectations. Furthermore, because 1 of these resulted from a genuine mutation, the error rate due to MDA is 7/1568 (0.45%). We conclude that many hundreds of microsatellites or other genetic markers can be accurately genotyped from a single miracidium using this method, greatly expanding the scope of population genetic, epidemiological and evolutionary studies on this parasite.
Valentim
Claudia L L
CL
Department of Genetics, Southwest Foundation for Biomedical Research, San Antonio, TX 78245, USA.
LoVerde
Philip T
PT
Anderson
Timothy J C
TJ
Criscione
Charles D
CD
D000818:Animals; D005260:Female; D005821:Genetic Techniques / Q000592:standards*; D049751:Genome, Helminth / Q000235:genetics*; D005838:Genotype; D008018:Life Cycle Stages; D008297:Male; D018895:Microsatellite Repeats / Q000235:genetics; D015203:Reproducibility of Results; D012550:Schistosoma mansoni / Q000235:genetics*