Direct reverse transcription--polymerase chain reaction from whole blood without RNA extraction.
A simple, fast, efficient method of direct reverse transcription-polymerase chain reaction (RT-PCR) from whole blood without RNA extraction is described. After the erythrocytes in a few microliters of fresh blood were destroyed and removed, the total RNA was released from the leukocytes by means of water treated with diethyl pyrocarbonate. The solution was used as RNA template and directly introduced in the RT-PCR. Following this method, which is called direct RT-PCR, three different cDNA fragments were successfully amplified with three different pairs of primers, respectively. Meanwhile, the direct RT-PCR was compared with the RT-PCR from the extracted total RNA template. The results showed no obvious difference.
Shi
Y J
YJ
Department of Biochemistry and Molecular Biology, Chinese Academy of Medical Sciences, Beijing.
Liu
J Z
JZ
D004277:DNA, Single-Stranded / Q000032:analysis; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D012313:RNA / Q000032:analysis* / Q000097:blood* / Q000302:isolation & purification; D012194:RNA-Directed DNA Polymerase; D012260:Ribonucleases / Q000037:antagonists & inhibitors
D004277:DNA, Single-Stranded; D012313:RNA; D012194:RNA-Directed DNA Polymerase; D012260:Ribonucleases