A single erythroid-specific DNase I super-hypersensitive site activates high levels of human beta-globin gene expression in transgenic mice.
A single erythroid-specific DNase I super-hypersensitive site activates high levels of human beta-globin gene expression in transgenic mice.
复制标题
单个红细胞特异性 DNase I 超敏感位点可激活转基因小鼠中高水平的人类 β-珠蛋白基因表达。
DOI：
10.1101/gad.3.3.314
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发表时间：
1989
影响因子：
10.5
通讯作者：
Brinster,RL
中科院分区：
文献类型：
--
作者：
Ryan,TM;Behringer,RR;Martin,NC;Townes,TM;Palmiter,RD;Brinster,RL
将通常位于人β-珠蛋白基因座远上游的红细胞特异性DNA酶I超敏（HS）位点直接插入含有人β-珠蛋白基因的4.1 kb片段的上游。将这些构建体（HS β）和仅含有β-珠蛋白基因的构建体（β）显微注射到受精的小鼠卵中，并在发育的第16天胚胎的红系胎肝和脑中分析表达。仅含有β基因的23只动物中只有7只在红系组织中表达人β-珠蛋白mRNA，每个基因拷贝的平均表达水平为内源性小鼠β-珠蛋白mRNA的0.3%。相比之下，51只含有各种HS β构建体的转基因小鼠中有50只在红系组织中特异性表达转基因。含有所有5个上游HS位点的构建体的每个基因拷贝的平均表达水平为内源性小鼠β-珠蛋白mRNA的109%。含有单个超敏位点（HS II β）的构建体表达的人β-珠蛋白是每个基因拷贝的小鼠β-珠蛋白mRNA的40%。这些结果表明，HS VI位点，通常位于下游的人β-珠蛋白基因座，是不需要高水平表达。此外，结果表明，即使当含有红细胞特异性超敏位点II（HS II）的相对小的DNA片段（1.9kb）插入人β-珠蛋白基因的上游时，也可以在转基因小鼠中获得高水平的人β-珠蛋白基因表达。
Erythroid-specific DNase I super-hypersensitive (HS) sites that are normally located far upstream of the human beta-globin locus were inserted immediately upstream of a 4.1-kb fragment containing the human beta-globin gene. These constructs (HS beta) and a construct containing the beta-globin gene alone (beta) were microinjected into fertilized mouse eggs, and expression was analyzed in erythroid fetal liver and brain of day-16 embryos that developed. Only 7 of 23 animals that contained the beta gene alone expressed human beta-globin mRNA in erythroid tissue, and the average level of expression per gene copy was 0.3% of endogenous mouse beta-globin mRNA. In contrast, 50 of 51 transgenic mice that contained various HS beta constructs expressed the transgene specifically in erythroid tissue. The average level of expression per gene copy for constructs containing all five upstream HS sites was 109% of endogenous mouse beta-globin mRNA. Constructs that contained a single super-hypersensitive site (HS II beta) expressed 40% as much human beta-globin as mouse beta-globin mRNA per gene copy. These results demonstrate that the HS VI site, normally located downstream of the human beta-globin locus, is not required for high-level expression. Furthermore, the results demonstrate that high levels of human beta-globin gene expression can be obtained in transgenic mice even when a relatively small fragment of DNA (1.9 kb) containing erythroid-specific super-hypersensitive site II (HS II) is inserted upstream of the human beta-globin gene.