Isolation and characterization of kinetoplast DNA networks and minicircles from Crithidia fasciculata.
Isolation and characterization of kinetoplast DNA networks and minicircles from Crithidia fasciculata.
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Crithidia fasciculata 动质体 DNA 网络和小环的分离和表征。
DOI：
10.1111/j.1550-7408.1974.tb03751.x
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发表时间：
1974
期刊：
影响因子：
--
通讯作者：
L. Simpson
中科院分区：
文献类型：
--
作者：
A. M. Simpson;L. Simpson
概要。通过在低离心场选择性沉淀网络的技术，已从静止期 Crithidia fasciculata 细胞中分离出共价闭合动质体 DNA 网络。通过简单的差速离心，回收了大约 62% 的细胞动质体 DNA，不含核 DNA。
纯化的动质体 DNA 网络在电子显微镜和光学显微镜下均可见。在中性和碱性蔗糖中，闭合网络沉积为均匀带，中性蔗糖中的 s20w 约为 5 × 103。
通过在碱性蔗糖中温和超声处理和条带沉降，从纯化的网络中分离出闭合的单体小环。测量了闭合单体微环的几种物理性质。其中包括分子量、CsCl 中的浮力密度、超螺旋密度和沉降系数。
SYNOPSIS. Covalently closed kinetoplast DNA networks have been isolated from stationary phase Crithidia fasciculata cells by a technic involving selective pelleting of the networks at a low centrifugal field. Approximately 62% of the kinetoplast DNA of the cell was recovered free of nuclear DNA by simple differential centrifugation.
Purified kinetoplast DNA networks were visualized both in the electron microscope and in the light microscope. Closed networks sedimented as a homogeneous band both in neutral and alkaline sucrose, with an s20w in neutral sucrose of approximately 5 × 103.
Closed monomeric minicircles were isolated from purified networks by mild sonication and band sedimentation in alkaline sucrose. Several physical properties of closed monomeric minicircles were measured. These included molecular weight, buoyant density in CsCl, superhelix density and sedimentation coefficient.