IMPROVED MAMMALIAN VECTORS FOR HIGH EXPRESSION OF G418 RESISTANCE
IMPROVED MAMMALIAN VECTORS FOR HIGH EXPRESSION OF G418 RESISTANCE
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DOI：
10.1247/csf.12.575
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发表时间：
1987-12-01
影响因子：
1.5
通讯作者：
KONDOH, H
中科院分区：
文献类型：
--
作者：
KATOH, K;TAKAHASHI, Y;KONDOH, H
细胞系，例如畸胎瘤细胞和胚胎干细胞的细胞系，在转染neo载体后未能支持高G418抗性。为了减轻这种情况，我们分两步修饰pSV 2-neo，首先用SV 40早期基因和HSVtk的串联启动子，然后通过去除位于mRNA序列中真实起始密码子上游的不适当的met密码子。在最终产物pSTneoB中，neo转录单位在两端具有XhoI位点。pSTneoB以比pSV 2-neo显著更高的效率产生畸胎瘤细胞的G418抗性转化体。该载体将G418选择在基因转移中的应用扩展到更广泛的细胞类型。
Cell lines, such as those of teratocarcinoma an embryonic stem cells, fail to support high G418 resistance after transfection of neo vectors. To alleviate this, we modified pSV2-neo in two steps, first with tandem promoters of SV40 early genes and HSVtk, then by removing an improper met codon located immediately upstream of the authentic initiator codon in the mRNA sequence. In the final product, pSTneoB, the neo transcription unit has XhoI sites at both ends. pSTneoB yields G418-resistant transformants of teratocarcinoma cells at dramatically higher efficiencies than pSV2-neo. This vector extends the application of G418 selection in gene transfer to a wider range of cell types.