CLONING AND CHARACTERIZATION OF A COMPLEX SATELLITE DNA FROM DROSOPHILA-MELANOGASTER
CLONING AND CHARACTERIZATION OF A COMPLEX SATELLITE DNA FROM DROSOPHILA-MELANOGASTER
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DOI：
10.1016/0092-8674(77)90054-x
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发表时间：
1977-01-01
期刊：
影响因子：
64.5
通讯作者：
BRUTLAG, D
中科院分区：
文献类型：
--
作者：
CARLSON, M;BRUTLAG, D
对1.688卫星DNA（CsCl中密度为1.688g/cm ~ 3）的序列结构进行了分析，发现该卫星DNA与其它D.黑腹鱼卫星DNA具有更大的序列复杂性。通过连续的平衡密度离心纯化1.688卫星DNA，得到纯度为77%的级分。通过在质粒载体pSC 101中的分子克隆分离卫星DNA片段。其中一个重组质粒含有1.688个卫星DNA片段，大小为5.8个酶对，在大肠杆菌中繁殖过程中稳定。埃及嗜血杆菌（Hae III）、H.流感f（Hinf）和藤黄节杆菌（Alu I）定位在该杂合质粒的卫星DNA中。Hae III、Hinf和2个Alu I位点以约365个碱基对的规则间隔排列，有力地证明了该卫星DNA的序列复杂性为365个碱基对。进一步的证据来自发现，梯度纯化和克隆的1.688卫星DNA复性与它们的Hae II位点对齐。梯度纯化的卫星DNA中的Hae III和Hinf位点被其他人显示为以365个碱基对及其整数倍的间隔分布。在其他规则阵列中的一些位点被随机灭活。克隆的卫星DNA提供了一个杂交探针的梯度纯化的卫星DNA中的这些识别位点的安排的敏感性研究。卫星DNA的某些区域包含的识别位点比所提出的模型所预期的要少得多。1.688卫星DNA的不同区域可能表现出不同的Hae III和Hinf识别位点的排列。
The sequence organization of the 1.688 satellite DNA (density 1.688 g/cm3 in CsCl) were investigated, and this satellite differed from the other D. melanogaster satellite DNA in having a much greater sequence complexity. Purification of 1.688 satellite DNA by successive equilibrium density centrifugations yielded a fraction 77% pure. Segments of satellite DNA were isolated by molecular cloning in the plasmid vector pSC101. One recombinant plasmid contained a segment of 1.688 satellite DNA 5.8 kilobase pairs in size and was stable during propagation in Escherichia coli. Recognition sites for restriction enzymes from Haemophilus aegyptius (Hae III), H. influenzae f (Hinf) and Arthrobacter luteus (Alu I) were mapped in the satellite DNA of this hybrid plasmid. The spacing of Hae III, Hinf and 2 Alu I sites at regular intervals of about 365 base pairs is strong evidence that the sequence complexity of this satellite DNA is 365 base pairs. Further evidence comes from the finding that both gradient-purified and cloned 1.688 satellite DNA renature with their Hae II sites in register. The Hae III and Hinf sites in gradient-purified satellite DNA were shown by others to be distributed at intervals of 365 base pairs and integral multiples thereof. Some of the sites in an otherwise regular array were randomly inactivated. Cloned satellite DNA provided a hybridization probe for sensitive studies of the arrangement of these recognition sites in gradient-purified satellite DNA. Some regions of satellite DNA contained many fewer recognition sites than expected from the proposed models. Different regions of 1.688 satellite DNA may exhibit different arrangements of Hae III and Hinf recognition sites.