RUN-OFF SYNTHESIS AND APPLICATION OF DEFINED SINGLE-STRANDED-DNA HYBRIDIZATION PROBES

被引:55
作者
STURZL, M
ROTH, WK
机构
[1] Department of Virus Research, Max-Planck-Institut für Biochemie
关键词
D O I
10.1016/0003-2697(90)90274-D
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
A simple and efficient method for synthesizing radioactively labeled single-stranded DNA hybridization probes with Thermus aquaticus (Taq) DNA polymerase is described. This is done in a "run-off" polymerization with repeated cycles of denaturation, annealing, and extension. It leads to high yields of a single-stranded DNA of defined length (up to 5000 nt), which is labeled to a high specific activity (1.3 × 108 cpm/μg DNA). These hybridization probes are equally sensitive as nick-translated DNA probes, but strand specific. This was tested by slot blot hybridization with in vitro-transcribed target RNAs and by Northern blotting. The use of single-stranded DNA hybridization probes combines the benefits of DNA stability and single-strand RNA probes. © 1990.
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收藏
页码:164 / 169
页数:6
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