A supersensitive dot-hybridization method: rapid and quantitative detection of host-derived DNA in recombinant products at the one picogram level.
We have developed a highly sensitive method of DNA dot-blotting hybridization to detect host-derived DNA (nuclear DNA and plasmid DNA) in a recombinant product. This method has two distinctive features compared to the conventional hybridization method: firstly, a highly specific radioactive probe is prepared by using ultrasonicated DNA, instead of untreated DNA, as a template for the oligo-labeling reaction; secondly, the signal to noise ratio is increased by the use of lambda phage DNA as non-homologous DNA. This method enabled us to detect host-derived DNA at the one picogram level without using a radioisotope of high specific activity and long exposure times.
Kuroda
S
S
Central Research Division, Takeda Chemical Industries, Ltd., Osaka, Japan.
Itoh
Y
Y
Miyazaki
T
T
Fujisawa
Y
Y
D002467:Cell Nucleus / Q000032:analysis; D004271:DNA, Fungal / Q000032:analysis*; D004274:DNA, Recombinant / Q000032:analysis*; D005818:Genetic Engineering / Q000379:methods; D006514:Hepatitis B Surface Antigens / Q000235:genetics; D008832:Microchemistry; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D014176:Protein Biosynthesis; D012441:Saccharomyces cerevisiae / Q000235:genetics
D004271:DNA, Fungal; D004274:DNA, Recombinant; D006514:Hepatitis B Surface Antigens