Accurate processing and amplification of cloned germ line copies of ribosomal DNA injected into developing nuclei of Tetrahymena thermophila.
Accurate processing and amplification of cloned germ line copies of ribosomal DNA injected into developing nuclei of Tetrahymena thermophila.
复制标题
准确处理和扩增注射到嗜热四膜虫发育细胞核中的核糖体 DNA 的克隆种系拷贝。
DOI：
10.1128/mcb.9.3.1092-1099.1989
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发表时间：
1989
影响因子：
5.3
通讯作者：
Yao,CH
中科院分区：
文献类型：
--
作者：
Yao,MC;Yao,CH
纤毛虫四膜虫在其生发(微核)基因组中含有rRNA基因(RDNA)的染色体整合拷贝。这些基因通过涉及定点DNA断裂的过程从染色体上切除，通过增加端粒成为线性回文分子，并在体细胞核(大核)的发育过程中得到极大的扩增。在这项研究中，我们克隆了含有这些基因的胚系DNA的15kb碱基片段，并将其注入发育中的大核。嗜热菌。高达11%的注射细胞转化为由注射的DNA指定的帕罗霉素耐药表型。转化效率取决于注射细胞的发育阶段和注射的DNA的完整性，而不是DNA的浓度或构象。注射的DNA显然经过了正确的加工和扩增，产生了具有预期线性回文结构的rDNA分子，该结构携带了适当的物理标记。因此，15kb的DNA含有足以导致rDNA扩增int的DNA加工事件的所有顺式作用序列。嗜热菌。
The ciliateTetrahymena thermophilacontains a chromosomally integrated copy of the rRNA genes (rDNA) in its germinal (micronuclear) genome. These genes are excised from the chromosome through a process involving site-specific DNA breakage, become linear palindromic molecules with added telomeres, and are greatly amplified during development of the somatic nucleus (macronucleus). In this study, we cloned a 15-kilobase segment of the germ line DNA containing these genes and injected it into developing macronuclei ofT. thermophila. Up to 11% of injected cells were transformed to the paromomycin-resistant phenotype specified by the injected DNA. Transformation efficiency was dependent on the developmental stages of the injected cells and the integrity of the injected DNA but not the DNA concentration or conformation. The injected DNA was apparently processed and amplified correctly to produce rDNA molecules with the expected linear palindromic structure which carried the appropriate physical markers. Thus, the 15-kilobase DNA contained allcis-acting sequences sufficient for the DNA-processing events leading to rDNA amplification inT. thermophila.