Transformation of frog embryos with a rabbit fB-globin gene
Transformation of frog embryos with a rabbit fB-globin gene
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用兔 fB 珠蛋白基因转化青蛙胚胎
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通讯作者：
W. Schaffner
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作者：
S. Rusconi;W. Schaffner
为了研究外源DNA在非洲爪蟾胚胎发生过程中的命运和可能的表达，我们将兔f1-珠蛋白基因注入非洲爪蟾受精卵中。在不同发育阶段提取蛙胚胎DNA，通过琼脂糖凝胶电泳进行分级，转移到硝酸纤维素滤膜上，并与标记的P-珠蛋白重组质粒DNA杂交。结果发现，注入的DNA复制染色体外，达到，在原肠胚阶段，相当于输入DNA的10至200倍扩增的水平。在后期阶段，大部分外源DNA被降解，但保留了一小部分。六周大的蝌蚪和六个月大的青蛙每个细胞平均含有3-10个兔珠蛋白基因拷贝。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，并在琼脂糖凝胶电泳中与高分子量爪蟾DNA共迁移。通过SI核酸酶图谱分析珠蛋白基因表达表明，兔fl-珠蛋白启动子在青蛙胚胎中被识别，并且转录本被正确剪接。
In order to study the fate and possible expression of foreign DNA during embryogenesis ofthe frog Xenopus laevis, we have injected a rabbit fi-globin gene 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 P-globin recombinant plasmid DNA. It was found that 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. Six-week-old tadpoles as well as six-month-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 elec-trophoresis with high molecular weight Xenopus DNA. Analysis of globin gene expression by SI nuclease mapping showed that the rabbit fl-globin promoter was recognized in the frog embryo and that the transcripts were correctly spliced.