urine samples, regardless of whether it was obtained from male or
female donors, indicates that the proposed DNA extraction method
is widely applicable for a variety of urine samples, as has been
confirmed by our repeated experiments.
As the recovery of cells and the purification of DNA were all
accomplished with rapid magnetic separation rather than by centrifugal
processes, the present procedure required less handling
and could be carried out in a single microcentrifuge tube. Further,
the methodology described requires no hazardous reagents (such
as phenol and chloroform) during the entire process. Finally, the
entire CMNP extraction process required less than 30 min, a significant
time savings over the traditional phenol/chloroform method,
which needed at least several hours to complete [9,10].
4. Conclusion
Carboxylated magnetic nanoparticles were used to develop a
novel method for PCR-ready genomic DNA extraction from urine
samples. CMNPs were used to both extract cells from urine samples,
and adsorb genomic DNA from the lysates. Sedimentation associated
with the freezing of urine samples could be re-dissolved with
the addition of EDTA and a subsequent pH adjustment, allowing
cells to be re-suspended for highly efficient binding by magnetic
nanoparticles. The quality of extracted urine DNA was confirmed
by PCR amplification of different gene fragments of different sizes.
Relative to traditional methods, the present procedure required
less handling, no hazardous reagents, and could be carried out in a
single microcentrifuge tube within 30 min. These methods, while
simple, rapid, sensitive and environmentally friendly, are suitable
for routine laboratory use, but also hold promise for construction
of automated urine extraction systems for various diagnostic purposes.
Acknowledgments
This work was a project supported by Scientific Research
Fund of Sichuan Provincial Education Department and supported
by Double-support Plan of Sichuan Agricultural University. The
authors thank Feng Chen for helpful suggestions.