Kerem B S, Goitein R, Diamond G, Cedar H, Marcus M
Cell. 1984 Sep;38(2):493-9. doi: 10.1016/0092-8674(84)90504-x.
We have shown that in fixed mitotic chromosomes from female G. gerbillus cells the inactive X chromosome is distinctly less sensitive to DNAase I than the active X chromosome, as demonstrated by in situ nick translation. These results indicated that the specific chromatin conformation that renders potentially active genes sensitive to DNAase I is maintained in fixed mitotic chromosomes. We increased the sensitivity and accuracy of in situ nick translation using biotinylated dUTP and a specific detection and staining procedure instead of radioactive label and autoradiography and now show that in both human and CHO chromosomes, the DNAase I sensitive and insensitive chromosomal regions form a specific dark and light banding pattern. The DNAase I sensitive dark D-bands usually correspond to the light G-bands, but not all light G-bands are DNAase I sensitive. Identifiable regions of inactive constitutive heterochromatin are in a DNAase I insensitive conformation. Our methodology provides a new and important tool for studying the structural and functional organization of chromosomes.
我们已经证明，在雌性沙鼠细胞固定的有丝分裂染色体中，通过原位缺口平移法显示，失活的X染色体对DNA酶I的敏感性明显低于活性X染色体。这些结果表明，使潜在活性基因对DNA酶I敏感的特定染色质构象在固定的有丝分裂染色体中得以维持。我们使用生物素化的dUTP以及特定的检测和染色程序，而非放射性标记和放射自显影，提高了原位缺口平移法的灵敏度和准确性，现在表明在人类和中国仓鼠卵巢细胞（CHO）染色体中，DNA酶I敏感和不敏感的染色体区域形成了特定的暗带和亮带模式。DNA酶I敏感的暗D带通常对应于亮G带，但并非所有亮G带都对DNA酶I敏感。可识别的非活性组成型异染色质区域处于对DNA酶I不敏感的构象。我们的方法为研究染色体的结构和功能组织提供了一个新的重要工具。
