TRANSFORMATION OF FROG EMBRYOS WITH A RABBIT BETA-GLOBIN GENE
TRANSFORMATION OF FROG EMBRYOS WITH A RABBIT BETA-GLOBIN GENE
复制标题
DOI：
10.1073/pnas.78.8.5051
复制
发表时间：
1981-01-01
期刊：
影响因子：
--
通讯作者：
SCHAFFNER, W
中科院分区：
文献类型：
--
作者：
RUSCONI, S;SCHAFFNER, W
为了研究外源DNA在非洲爪蛙胚胎发生过程中的去向和可能的表达，将兔β-珠蛋白基因注入非洲爪蛙受精卵中。提取不同发育阶段的青蛙胚胎DNA，经琼脂糖凝胶电泳法分离，转移到硝酸纤维素膜上，与标记的β-珠蛋白重组质粒DNA杂交。注入的DNA在染色体外复制，在原肠胚期达到相当于输入DNA扩增10至200倍的水平。在后期，大部分外源DNA被降解，但一小部分保持不变。6周大的蝌蚪和6个月大的青蛙平均每个细胞含有3-10个兔珠蛋白基因。这些持续存在的珠蛋白基因大多以长串联重复序列的形式存在，并在高分子量非洲爪哇DNA琼脂糖凝胶电泳中迁移。用S1核酸酶图谱分析珠蛋白基因的表达，表明兔β-珠蛋白启动子在青蛙胚胎中被识别，转录本被正确剪接。
To study the fate and possible expression of foreign DNA during embryogenesis of the frog X. laevis, a rabbit .beta.-globin gene was injected into fertilized Xenopus eggs. Frog embryo DNA was extracted at various stages of development, fractionated by agarose gel electrophoresis, transferred to nitrocellulose filters, and hybridized to labeled .beta.-globin recombinant plasmid DNA. The injected DNA replicated extrachromosomally, reaching, at gastrula stage, a level equivalent to a 10- to 200-fold amplification of input DNA. At later stages, a majority of the foreign DNA was degraded, but a small fraction was maintained. The 6-wk-old tadpoles and 6-mo.-old frogs contained an average of 3-10 copies of the rabbit globin gene per cell. Most of these persisting globin genes were present as long tandem repeats and comigrated in agarose gel electrophoresis with high MW Xenopus DNA. Analysis of globin gene expression by S1 nuclease mapping showed that the rabbit .beta.-globin promoter was recognized in the frog embryo and that the transcripts were correctly spliced.