Comparison of six nucleic acid extraction methods for detection of viral DNA or RNA sequences in four different non-serum specimen types.
BACKGROUND
Extraction of viral nucleic acids from serum samples is widely used in diagnostic pathology tests. However, the heterogeneous nature of non-serum samples may contribute to variations in the yields of viral nucleic acids with different extraction methods and specimen types.


OBJECTIVE
Six different methods were compared for optimal extraction of viral DNA or RNA from four types of non-serum specimens.


METHODS
The DNA viruses used were herpes simplex virus and cytomegalovirus. The RNA viruses were poliovirus, rotavirus and small round structured virus. The specimens used were from respiratory, genital, faecal and peripheral blood mononuclear cell samples. The extracted nucleic acids were amplified by PCR and detected in an enzyme immunoassay using digoxygenin-labelled amplicons.


CONCLUSIONS
For extraction of viral DNA, the phenol-chloroform method yielded the highest amount of DNA as judged by endpoint titration. The three methods compared for extraction of viral RNA used guanidine isothiocyanate and the QiaRNA kit was shown to yield the highest amount of viral RNA.
Kok
T
T
Infectious Diseases Laboratories, Institute of Medical and Veterinary Science, Adelaide, Australia. tuckweng.kok@imvs.sa.gov.au
Wati
S
S
Bayly
B
B
Devonshire-Gill
D
D
Higgins
G
G
D003587:Cytomegalovirus / Q000235:genetics / Q000302:isolation & purification; D004279:DNA, Viral / Q000302:isolation & purification*; D005243:Feces / Q000821:virology; D005835:Genitalia / Q000821:virology; D006801:Humans; D008214:Lymphocytes / Q000821:virology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012328:RNA Viruses / Q000235:genetics / Q000302:isolation & purification; D012367:RNA, Viral / Q000302:isolation & purification*; D012137:Respiratory System / Q000821:virology; D018139:Simplexvirus / Q000235:genetics / Q000302:isolation & purification
D004279:DNA, Viral; D012367:RNA, Viral