Homogeneity and persistence of transgene expression by omitting antibiotic selection in cell line isolation.
Homogeneity and persistence of transgene expression by omitting antibiotic selection in cell line isolation.
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DOI：
10.1093/nar/gkn508
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发表时间：
2008-10
影响因子：
14.9
通讯作者：
Gossen, Manfred
中科院分区：
文献类型：
--
作者：
Kaufman, Weimin L.;Kocman, Ibrahim;Agrawal, Vishal;Rahn, Hans-Peter;Besser, Daniel;Gossen, Manfred
转基因的不均匀的马赛克表达模式通常与转录沉默有关，这是由外源DNA的表观遗传修饰引发的。这种表型在基因工程细胞和生物体中是常见现象。它们被广泛归因于转基因转录单位不同于内源基因的特征，使它们特别容易受到表观遗传下调的影响。与这一假设相反，我们表明用于分离稳定转染细胞的方法对转基因表达模式有最深远的影响。标准抗生素选择与细胞分选直接比较，建立稳定的细胞。只有后一种方法才能保证基因表达的高度均匀性和稳定性。对细胞分选步骤有用的标记基因大多编码荧光蛋白。然而，通过将这种方法与位点特异性重组相结合，它可以用于分离具有任何感兴趣基因所需表达特征的稳定细胞系。
Nonuniform, mosaic expression patterns of transgenes are often linked to transcriptional silencing, triggered by epigenetic modifications of the exogenous DNA. Such phenotypes are common phenomena in genetically engineered cells and organisms. They are widely attributed to features of transgenic transcription units distinct from endogenous genes, rendering them particularly susceptible to epigenetic downregulation. Contrary to this assumption we show that the method used for the isolation of stably transfected cells has the most profound impact on transgene expression patterns. Standard antibiotic selection was directly compared to cell sorting for the establishment of stable cells. Only the latter procedure could warrant a high degree of uniformity and stability in gene expression. Marker genes useful for the essential cell sorting step encode mostly fluorescent proteins. However, by combining this approach with site-specific recombination, it can be applied to isolate stable cell lines with the desired expression characteristics for any gene of interest.
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影响因子：
5.1
作者：
Chen, ZY;He, CY;Kay, MA
通讯作者：
Kay, MA
影响因子：
12.4
作者：
Chen, Zhi-Ying;Riu, Efren;Kay, Mark A.
通讯作者：
Kay, Mark A.
影响因子：
14.9
作者：
Gopalkrishnan, RV;Christiansen, KA;Fisher, PB
通讯作者：
Fisher, PB
影响因子：
46.9
作者：
Buchholz, F;Angrand, PO;Stewart, AF
通讯作者：
Stewart, AF
影响因子：
3.5
作者：
Migliaccio, AR;Bengra, C;Tuan, D
通讯作者：
Tuan, D