A novel buffer system, AnyDirect, can improve polymerase chain reaction from whole blood without DNA isolation.
BACKGROUND
Polymerase chain reaction (PCR) using DNA from blood samples is a valuable tool in the field of medical diagnostics. However, DNA isolation from blood is a laborious and sample-consuming step, and hampers the automation of PCR for large-scale studies. Attempts to perform PCR from blood without DNA isolation have been difficult to achieve, since numerous endogenous and exogenous blood constituents may inhibit PCR.


METHODS
We used a novel buffer system, 'AnyDirect', that conserves the enzymatic activity of DNA polymerases for effective use in direct PCR from whole blood under various conditions.


RESULTS
Using AnyDirect, DNA amplification was achieved from whole blood with a variety of thermostable DNA polymerases. Amplification occurred regardless of target size (up to 1.7 kb), presence of various known PCR inhibitors, and high target GC content. Importantly, low copy number DNA targets were effectively amplified from whole blood.


CONCLUSIONS
AnyDirect buffer allows direct PCR from whole blood and may facilitate detection of genetic diseases or infections by eliminating the time and effort for DNA extraction. The use of AnyDirect could facilitate the development of high-throughput PCR for large-scale diagnostic screening or investigation of various medical conditions.
Yang
Young Geun
YG
Department of Biochemistry, College of Science, Yonsei University, Seoul 120-749, Republic of Korea.
Kim
Jong Yeol
JY
Song
Young-Han
YH
Kim
Doo-Sik
DS
D002021:Buffers; D004247:DNA / Q000097:blood* / Q000737:chemistry; D017931:DNA Primers; D015497:HIV-1 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007202:Indicators and Reagents; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D019914:Taq Polymerase / Q000037:antagonists & inhibitors / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
D002021:Buffers; D017931:DNA Primers; D007202:Indicators and Reagents; D016159:Tumor Suppressor Protein p53; D004247:DNA; D019914:Taq Polymerase