HIGH-YIELD AND STABLE TRANSFORMATION OF THE UNICELLULAR GREEN-ALGA ACETABULARIA BY MICROINJECTION OF SV40 DNA AND PSV2NEO
HIGH-YIELD AND STABLE TRANSFORMATION OF THE UNICELLULAR GREEN-ALGA ACETABULARIA BY MICROINJECTION OF SV40 DNA AND PSV2NEO
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DOI：
10.1002/j.1460-2075.1986.tb04380.x
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发表时间：
1986-07-01
期刊：
影响因子：
11.4
通讯作者：
SCHWEIGER, HG
中科院分区：
文献类型：
--
作者：
NEUHAUS, G;NEUHAUSURL, G;SCHWEIGER, HG
将SV40 DNA和pSV2neo微量注射到地中海髋臼离体细胞核中。将注射的细胞核植入同种无核细胞片段中。这样的组合不仅存活了下来，而且还形成了后代。对这些组合的F1、F2和F3代进行分析。间接免疫荧光显示，在sv40处理的细胞中，t抗原在三代细胞的细胞核中均有表达和积累。核交换实验显示，t抗原的表达仅在细胞核转化时发生，而在细胞质转化时则不发生。pSV2neo是一种嵌合基因，具有选择性标记物，通过诱导G-418抗性和免疫荧光证实了其转化。从F1、F2和F3代注射细胞囊中的配子中分离基因组DNA，并进行Southern分析。这些实验表明，这两种类型的DNA都被整合到宿主基因组中。
SV40 DNA and pSV2neo were microinjected into isolated nuclei of Acetabularia mediterranea. The injected nuclei were implanted into anucleate cell fragments of the same species. Such combinations not only survived but also formed progeny. The F1, F2 and F3 generations of these combinations were analyzed. In the case of SV40-treated cells T-antigen was expressed and accumulated in the nuclei of all three generations studied as shown by indirect immunofluorescence. Nuclear exchange experiments revealed expression of the T-antigen only if a transformed nucleus but not if only a transformed cytoplasm was involved. Transformation by pSV2neo, a chimeric gene with a selectable marker was demonstrated by the induction of G-418 resistance as well as immunofluorescence. Genomic DNA was isolated from gametes, originating in cysts from the F1, F2 and F3 generations of injection cells, and subjected to Southern analysis. These experiments demonstrated that both types of DNA are integrated into the host genome.