Shchepinov M S, Udalova I A, Bridgman A J, Southern E M
Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, UK.
Nucleic Acids Res. 1997 Nov 15;25(22):4447-54. doi: 10.1093/nar/25.22.4447.
Oligonucleotide dendrimers were synthesized using a novel phosphoramidite synthon, tris-2,2,2-[3-(4,4'-dimethoxytrityloxy) propyloxymethyl]ethyl- N , N -diisopropylaminocyanethoxy phosphoramidite. Label, incorporated using [gamma-32P]ATP and polynucleotide kinase, was increased in proportion to the number of 5'-ends. There was a similar increase in signal when these multiply labelled oligonucleotides were used as probes to oligonucleotide arrays. A dendrimeric oligonucleotide was used successfully as a primer in the PCR. The strand bearing the dendrimer was resistant to degradation by T7 Gene 6 exonuclease making it easy to convert the double-stranded product of the PCR to a multiply-labelled, single-stranded probe.
使用一种新型亚磷酰胺合成子，即三-2,2,2-[3-(4,4'-二甲氧基三苯甲基氧基)丙氧基甲基]乙基-N,N-二异丙基氨基氰乙氧基亚磷酰胺，合成了寡核苷酸树枝状大分子。使用[γ-32P]ATP和多核苷酸激酶掺入的标记与5'-末端的数量成比例增加。当这些多重标记的寡核苷酸用作寡核苷酸阵列的探针时，信号也有类似的增加。一种树枝状寡核苷酸成功地用作PCR的引物。带有树枝状大分子的链对T7基因6核酸外切酶的降解具有抗性，这使得将PCR的双链产物转化为多重标记的单链探针变得容易。
