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首页> 外文期刊>Genesis: the journal of genetics and development >A retained selection cassette increases reporter gene expression without affecting tissue distribution in SPI3 knockout/GFP knock-in mice
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A retained selection cassette increases reporter gene expression without affecting tissue distribution in SPI3 knockout/GFP knock-in mice

机译:保留的选择盒可增加报告基因的表达,而不会影响SPI3基因敲除/ GFP基因敲入小鼠的组织分布

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摘要

The human serpin, proteinase inhibitor 6 (PI6/SERPINB6), is a protease inhibitor expressed in many tissues. It inhibits a large number of proteases, including cathepsin G in granulocytes and monocytes. To determine the temporal and spatial distribution of PI-6, mice were generated in which exon 2 of the PI-6 ortholog SPI3 (Serpinb6) was replaced with a green fluorescent protein (GFP) reporter gene. This placed GFP under the control of the regulatory elements and initiation codon of the SPI3 gene. The neomycin selection cassette was flanked by IoxP sites to allow excision from the targeted allele. GFP expression in heterozygous and SPI3-deficient mice accurately reflected the tissue distribution of SPI3 in all organs tested and allowed precise comparisons of expression levels. Interestingly, retention of the neomycin cassette in targeted mice resulted in 2-10-fold increases of GFP in leukocytes, but without affecting tissue-specific expression patterns. This is the first example of selection cassette retention specifically increasing reporter gene expression in targeted mice and reinforces the view that selection cassettes must be removed to avoid confounding effects on reporter gene expression patterns. genesis 36:149-157, 2003.
机译:人丝氨酸蛋白酶抑制剂蛋白酶抑制剂6(PI6 / SERPINB6)是在许多组织中表达的蛋白酶抑制剂。它抑制多种蛋白酶,包括粒细胞和单核细胞中的组织蛋白酶G。为了确定PI-6的时间和空间分布,产生了小鼠,其中用绿色荧光蛋白(GFP)报道基因替换了PI-6直向同源物SPI3(Serpinb6)的外显子2。这将GFP置于SPI3基因的调控元件和起始密码子的控制之下。新霉素选择盒的侧翼是IoxP位点,以允许从靶向等位基因上切除。 GFP在杂合子和SPI3缺陷型小鼠中的表达准确反映了SPI3在所有测试器官中的组织分布,并允许精确比较表达水平。有趣的是,新霉素盒在靶标小鼠中的保留导致白细胞中GFP的2-10倍增加,但不影响组织特异性表达模式。这是选择盒保留的第一个例子,其特异性地增加了靶向小鼠中报道基因的表达,并增强了必须去除选择盒以避免对报道基因表达模式造成混淆的观点。创世纪36:149-157,2003。

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