EFFICIENT GENE-TRANSFER IN C-ELEGANS - EXTRACHROMOSOMAL MAINTENANCE AND INTEGRATION OF TRANSFORMING SEQUENCES
EFFICIENT GENE-TRANSFER IN C-ELEGANS - EXTRACHROMOSOMAL MAINTENANCE AND INTEGRATION OF TRANSFORMING SEQUENCES
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DOI：
10.1002/j.1460-2075.1991.tb04966.x
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发表时间：
1991-12-01
期刊：
影响因子：
11.4
通讯作者：
AMBROS, V
中科院分区：
文献类型：
--
作者：
MELLO, CC;KRAMER, JM;AMBROS, V
我们描述了一个显性的行为标记，rol-6（苏-1006），和一个有效的显微注射程序，促进秀丽隐杆线虫转化体的恢复。我们利用这些工具来研究C.线虫DNA转化。通过注射遗传标记的DNA分子的混合物，我们表明，大的染色体外阵列组装直接从注射的分子和同源重组驱动阵列组装。适当放置的双链断裂刺激同源重组阵列形成过程中。我们的数据表明，组装的转基因结构的大小决定了它们是否将保持染色体外或丢失。我们表明，低拷贝数的染色体外转化可以通过调整注射混合物中DNA分子的相对浓度来实现。当单链寡核苷酸与双链DNA共注射时，注射的DNA的整合虽然相对罕见，但可再现地实现。
We describe a dominant behavioral marker, rol-6(su-1006), and an efficient microinjection procedure which facilitate the recovery of Caenorhabditis elegans transformants. We use these tools to study the mechanism of C. elegans DNA transformation. By injecting mixtures of genetically marked DNA molecules, we show that large extrachromosomal arrays assemble directly from the injected molecules and that homologous recombination drives array assembly. Appropriately placed double-strand breaks stimulated homologous recombination during array formation. Our data indicate that the size of the assembled transgenic structures determines whether or not they will be maintained extrachromosomally or lost. We show that low copy number extrachromosomal transformation can be achieved by adjusting the relative concentration of DNA molecules in the injection mixture. Integration of the injected DNA, though relatively rare, was reproducibly achieved when single-stranded oligonucleotide was co-injected with the double-stranded DNA.