Generation of albino Xenopus tropicalis using zinc-finger nucleases
Generation of albino Xenopus tropicalis using zinc-finger nucleases
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DOI：
10.1111/dgd.12006
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发表时间：
2012-12-01
影响因子：
2.5
通讯作者：
Yaoita, Yoshio
中科院分区：
文献类型：
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作者：
Nakajima, Keisuke;Nakajima, Taeko;Yaoita, Yoshio
为了产生热带爪蟾的白化病品系，我们将编码锌指核酸酶（ZFN）的mRNA注射到受精卵中，靶向酪氨酸酶编码区。令人惊讶的是，在注射了ZFN mRNA的F0青蛙的皮肤上观察到白癜风，表明基因组中的两个酪氨酸酶基因在白癜风斑块内的所有黑素细胞中被破坏。使用基因组DNA从皮肤突变分析显示，两个马赛克F0青蛙进行空间复杂的酪氨酸酶基因突变。这些数据表明，ZFN诱导的酪氨酸酶基因消融在整个身体的空间和时间上随机发生，可能直到幼蝌蚪阶段，并且缺乏功能性酪氨酸酶的黑素细胞前体增殖并形成白癜风斑块。几种白化病X.通过使嵌合F0青蛙交配，获得了双等位基因酪氨酸酶突变的复合杂合子tropicalis。据我们所知，这是第一次报道的白化病脊椎动物产生的靶向基因敲除。
To generate albino lines of Xenopus tropicalis, we injected fertilized eggs with mRNAs encoding zinc-finger nucleases (ZFNs) targeting the tyrosinase coding region. Surprisingly, vitiligo was observed on the skin of F0 frogs that had been injected with ZFN mRNAs, indicating that both tyrosinase genes in the genome were disrupted in all melanocytes within the vitiligo patches. Mutation analysis using genomic DNA from the skin revealed that two mosaic F0 frogs underwent spatially complex tyrosinase gene mutations. The data implies that the ZFN-induced tyrosinase gene ablations occurred randomly over space and time throughout the entire body, possibly until the young tadpole stage, and that melanocyte precursors lacking functional tyrosinase proliferated and formed vitiligo patches. Several albino X. tropicalis, which are compound heterozygotes for biallelic tyrosinase mutations, were obtained by mating the mosaic F0 frogs. To our knowledge, this is the first report of the albino vertebrates generated by the targeted gene knockout.