General protocol for microcell-mediated chromosome transfer
General protocol for microcell-mediated chromosome transfer
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微细胞介导的染色体转移的通用方案
DOI：
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发表时间：
1987
期刊：
影响因子：
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通讯作者：
E. Stubblefield
中科院分区：
文献类型：
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作者：
J. Sanford;E. Stubblefield
我们已经开发了一种通用的技术，使微核细胞用于微细胞介导的染色体转移。用秋水仙胺阻断生长细胞的有丝分裂，置于低渗溶液中10分钟，并返回培养基24小时。该处理促进淋巴母细胞或成纤维细胞内微核的形成。微细胞是通过在Percoll密度梯度上以43，500 g离心的细胞松弛素B处理产生的。将所得微细胞、全细胞和有核体的混合物通过3 μm孔过滤，以获得纯微细胞制剂。使用植物血凝素-P和聚乙二醇将微细胞与受体全细胞融合。这种技术的优点是：（1）供体细胞不需要附着在基质上;（2）低频率形成微核的细胞系仍然可以有效地用作微细胞供体。
We have developed a general technique for making micronucleated cells to use in microcell-mediated chromosome transfer. Growing cells are blocked in mitosis with colcemid, placed in a hypotonic solution for 10 min, and returned to culture medium for 24 h. This treatment promotes the formation of micronuclei within lymphoblast or fibroblast cells. The microcells are generated by cytochalasin B treatment on a Percoll density gradient centrifuged at 43,500g. The resulting mixture of microcells, whole cells, and karyoplasts is filtered through 3-μm pores to obtain a pure microcell preparation. The microcells are fused to recipient whole cells using phytohemagglutinin-P and polyethylene glycol. Advantages of this technique are: (1) donor cells need not be attached to a substrate; and (2) cell lines which form micronuclei in low frequency can still be used efficiently as microcell donors.