Köster H, Albersmeyer K, Skroch D
Hoppe Seylers Z Physiol Chem. 1978 Nov;359(11):1579-89. doi: 10.1515/bchm2.1978.359.2.1579.
Two methods are described by which primer molecules like UpU and oligodeoxythymidylates can be coupled with high efficiency to an insoluble polymer, like hydroxypropylated Sephadex G-50, by one covalent linkage. In one procedure aliphatic dicarboxylic dichlorides (e.g. adipoyl dichloride) are used to serve as spacers of variable length and for anchoring the primer molecule UpU. The other method involves pU as an anchor for (pdT)3 and (pdT)6, which are coupled to the polymer using condensation reactions with 2,4,6-triisopropylphenylsulfonyl chloride. In both cases the homogeneous primer molecules are bound specifically to the polymer. The insoluble primers are tested for their priming efficiency using polynucleotide nucleotidyltransferase from Micrococcus luteus and DNA nucleotidylexotransferase from calf thymus. The primers and synthesized polynucleotides can be cleaved from the polymer under conditions which are not damaging to ribo- and deoxyribopolynucleotides.
本文描述了两种方法，通过这两种方法，像UpU和寡聚脱氧胸苷酸这样的引物分子可以通过一个共价键高效地与不溶性聚合物（如羟丙基化葡聚糖凝胶G-50）偶联。在一种方法中，脂肪族二羧酸二氯化物（如己二酰二氯）用作可变长度的间隔物，并用于锚定引物分子UpU。另一种方法涉及使用pU作为（pdT）3和（pdT）6的锚定物，它们通过与2,4,6-三异丙基苯磺酰氯的缩合反应与聚合物偶联。在这两种情况下，均相引物分子都特异性地结合到聚合物上。使用来自藤黄微球菌的多核苷酸核苷酸转移酶和来自小牛胸腺的DNA核苷酸外转移酶测试不溶性引物的引发效率。引物和合成的多核苷酸可以在不损害核糖和脱氧核糖多核苷酸的条件下从聚合物上切割下来。
