A salting out and resin procedure for extracting Schistosoma mansoni DNA from human urine samples
1 Laboratório de Esquistossomose - Centro de Pesquisas René Rachou (CPqRR) - Fundação Oswaldo Cruz (FIOCRUZ), Av. Augusto de Lima 1715, Belo Horizonte, Minas Gerais, 30190-002, Brazil
2 Laboratório de Imunologia Celular e Molecular - Centro de Pesquisas René Rachou (CPqRR) - Fundação Oswaldo Cruz (FIOCRUZ), Av. Augusto de Lima 1715, Belo Horizonte, Minas Gerais, 30190-002, Brazil
3 Laboratório de Pesquisas Clínicas - Escola de Farmácia, Universidade Federal de Ouro Preto (UFOP), Campus Morro de Cruzeiro, Ouro Preto, Minas Gerais, 35400-00, Brazil
BMC Research Notes 2010, 3:115 doi:10.1186/1756-0500-3-115Published: 26 April 2010
In this paper a simple and cheap salting out and resin (InstaGene matrix® resin - BioRad) DNA extraction method from urine for PCR assays is introduced. The DNA of the fluke Schistosoma mansoni was chosen as the target since schistosomiasis lacks a suitable diagnostic tool which is sensitive enough to detect low worm burden. It is well known that the PCR technique provides high sensitivity and specificity in detecting parasite DNA. Therefore it is of paramount importance to take advantage of its excellent performance by providing a simple to handle and reliable DNA extraction procedure, which permits the diagnosis of the disease in easily obtainable urine samples.
The description of the extraction procedure is given. This extraction procedure was tested for reproducibility and efficiency in artificially contaminated human urine samples. The reproducibility reached 100%, showing positive results in 5 assay repetitions of 5 tested samples each containing 20 ng DNA/5 ml. The efficiency of the extraction procedure was also evaluated in a serial dilution of the original 20 ng DNA/5 ml sample. Detectable DNA was extracted when it was at a concentration of 1.28 pg DNA/mL, revealing the high efficiency of this procedure.
This methodology represents a promising tool for schistosomiasis diagnosis utilizing a bio-molecular technique in urine samples which is now ready to be tested under field conditions and may be applicable to the diagnosis of other parasitic diseases.