Eggert H, Jack R S
Institut für Genetik, Universität Köln, Germany.
EMBO J. 1991 May;10(5):1237-43. doi: 10.1002/j.1460-2075.1991.tb08065.x.
In vitro assays using detergent extracted nuclei allow the operational definition of SARs--specific sequences in the chromosome which are thought to interact with a structural matrix or scaffold. This interaction results in the formation of large stable protein-DNA complexes. We have used P-element transformation to introduce a characterized SAR into the Drosophila genome. The standard assay, which uses detergent extracted nuclei, shows that the ectopic SAR is indeed bound to the scaffold. However, in unextracted nuclei, a chromatin fragment containing the SAR sequence is eluted from the nucleus as readily as a fragment which lacks an SAR. Furthermore, an analysis of the accessibility of the neighbouring chromosomal restriction sites in unextracted nuclei indicates that the introduction of this ectopic SAR does not influence the local structure of chromatin. We conclude that the ectopic SAR site is not attached to a nuclear scaffold or matrix in vivo.
使用去污剂提取细胞核进行的体外测定使得能够对SARs(染色体中被认为与结构基质或支架相互作用的特定序列)进行操作性定义。这种相互作用导致形成大的稳定蛋白质 - DNA复合物。我们已利用P因子转化将一个已鉴定的SAR引入果蝇基因组。使用去污剂提取细胞核的标准测定表明,异位SAR确实与支架结合。然而,在未提取细胞核的情况下,含有SAR序列的染色质片段与缺乏SAR的片段一样容易从细胞核中洗脱出来。此外,对未提取细胞核中相邻染色体限制位点可及性的分析表明,这种异位SAR的引入并不影响染色质的局部结构。我们得出结论,异位SAR位点在体内未附着于核支架或基质。