【摘要】 从流感嗜血杆菌(Haemophilus influenzae)Rd 株中纯化了限制性内切核酸酶HindⅢ及部份纯化了Hind Ⅱ;探讨了纯化过程中的某些问题;检查了酶对底物双链DNA 的特异性内切作用,从琼脂糖平板凝胶电泳中观察到的经消化后产生的DNA 断片表明:酶对各种底物具有专一性作用;通过DNA 断片的重新连接证明:经Hind Ⅲ消化后生成的断片具有粘性末端,也说明纯化的Hind Ⅲ中不合明显的外切酶活力;用聚丙烯酰胺-十二烷基硫酸钠平板凝胶电泳,葡聚糖凝胶G150分子筛层析及蔗糖密度梯度离心法测定了Hind Ⅲ的分子量并对其亚单位进行了讨论.更多还原

【Abstract】 Hind Ⅲ and Hind Ⅱ have been purified from Haemophilus influenzae Rd strain.Some problems in the purification procedure were investigated.Endonucleolyticdigestion of DNA from plasmid pBR322 and phage λcI857S7 has been examined andthe DNA fragments resolved by agarose electrophoresis.The digestion profilesproduced are unique and characteristic for both DNA’s and the endonucleases.The ligation of Hind Ⅲ generated fragments indicates the existence of cohesiveends of the Hind Ⅲ products.It also shows that the preparation of Hind Ⅲis free of significant exonuclease activity.The molecular weight of Hind Ⅲ wasestimated to be 135,000 as determined by polyacrylamide gel electrophoresis,Seph-adex G150 gel filtration,and sucrose density gradient centrifugation.Electro-phoresis in the presence of SDS revealed two polypeptides,MW 70,000 and 62,000,which is consistent with the presence of nuclease and methylase componentsin this enzyme.更多还原