Werner D, Hadjiolov D, Hershey H V
Chem Biol Interact. 1981 Nov;37(3):279-87. doi: 10.1016/0009-2797(81)90114-9.
DNA released from Ehrlich ascites cells by lysis in the presence of 50 microgram X ml-1 of proteinase K contains long alkali-stable strands in the order of 50-100 X 10(6) daltons. In contrast, DNA released in the presence of 6 mg X ml-1 of autodigested pronase is significantly nicked. According to sedimentation rates the number of internal ends liberated during this procedure is 24/200 X 10(6) daltons. The number of alkali-labile sites introduced into DNA by incubation of Ehrlich ascites cells with 1 nM of N-methyl-N-nitrosourea (MNU) followed by cell lysis in the presence of 50 microgram X ml-1 of proteinase K and alkali-denaturation is 16.6/200 X 10(6) daltons. From this one should expect that denatured DNA released from cells pretreated with 1 mM of MNU which are subsequently lysed with 6 mg X ml-1 of pronase would have about 40 single-strand breaks/200 X 10(6) daltons. However, denatured DNA strands released by 6 mg X ml-1 of pronase either from MNU-treated or untreated cells cannot be separated by centrifugation through alkaline sucrose gradients. This phenomenon could be explained by a non-random distribution of MNU-inducible alkali-labile sites of DNA in vivo.
在含有50微克/毫升蛋白酶K的情况下通过裂解从艾氏腹水细胞释放的DNA含有长度约为50 - 100×10⁶道尔顿的长碱稳定链。相比之下，在含有6毫克/毫升自消化链霉蛋白酶的情况下释放的DNA有明显的切口。根据沉降速率，在此过程中释放的内部末端数量为24/200×10⁶道尔顿。将艾氏腹水细胞与1纳摩尔N - 甲基 - N - 亚硝基脲（MNU）孵育，随后在含有50微克/毫升蛋白酶K的情况下进行细胞裂解并碱变性，引入到DNA中的碱不稳定位点数量为16.6/200×10⁶道尔顿。由此可以预期，用1毫摩尔MNU预处理后再用6毫克/毫升链霉蛋白酶裂解的细胞释放的变性DNA每200×10⁶道尔顿大约会有40个单链断裂。然而，无论是来自经MNU处理还是未处理的细胞，6毫克/毫升链霉蛋白酶释放的变性DNA链都不能通过碱性蔗糖梯度离心分离。这种现象可以用体内DNA中MNU诱导的碱不稳定位点的非随机分布来解释。
