Observations on polymerase chain reaction amplification of infectious bursal disease virus dsRNA.
Two methods for denaturing double stranded (ds)RNA of infectious bursal disease virus for the purpose of reverse transcribing it were compared: Heat denaturation at 65 degrees C in the presence of DMSO and in the absence of DMSO. As part of the analysis, the nature of cDNA in the two preparations was examined by polymerase chain reaction (PCR) amplification, firstly by varying the number of cycles of PCR, and secondly by re-amplification of serial dilutions of the reaction products. The results show that denaturation of dsRNA in the presence of DMSO (method 1) is superior to denaturation without DMSO (method 2) judging by the yield of a specific PCR fragment after 30 cycles, and that the products of method 2 can be re-amplified, albeit poorly, with the generation of heterologous products.
Qian
B
B
Atlantic Veterinary College, University of Prince Edward Island Charlottetown, Canada.
Kibenge
F S
FS
D001483:Base Sequence; D004121:Dimethyl Sulfoxide; D007243:Infectious bursal disease virus / Q000235:genetics*; D008969:Molecular Sequence Data; D009691:Nucleic Acid Denaturation; D016133:Polymerase Chain Reaction / Q000379:methods*; D012330:RNA, Double-Stranded / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics*
D012330:RNA, Double-Stranded; D012367:RNA, Viral; D004121:Dimethyl Sulfoxide