REPLICATION, INTEGRATION AND EXPRESSION OF EXOGENOUS DNA INJECTED INTO FERTILIZED-EGGS OF XENOPUS-LAEVIS
REPLICATION, INTEGRATION AND EXPRESSION OF EXOGENOUS DNA INJECTED INTO FERTILIZED-EGGS OF XENOPUS-LAEVIS
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DOI：
10.1111/j.1432-0436.1984.tb01395.x
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发表时间：
1984-01-01
期刊：
影响因子：
2.9
通讯作者：
BEKTESH, SL
中科院分区：
文献类型：
--
作者：
ETKIN, LD;PEARMAN, B;BEKTESH, SL
分析了非洲爪哇受精卵显微注射后环状和线状DNA分子的去向。含有海胆组蛋白基因(PSP 102)、果蝇ADH基因(SAC-1)和SV40(SV2 CAT)的重组质粒在注射的青蛙胚胎发育过程中进行复制。相比之下，pBR322无论是作为单体还是作为多聚体，似乎都没有那么有效地复制。注射的环状DNA在原肠胚发育阶段是检测不到的，尽管有几个例子表明这些分子一直持续到幼虫阶段。在90%的病例中，注射的线性DNA以离散分子的形式持续存在到早期胚胎阶段。检测到与注入的线性和环状分子互补的DNA序列的一部分，从卵裂中期开始与高分子量细胞蛙DNA(48kb[kb]或更大)共游走。对注射的胚胎进行的DNA限制性内切酶分析表明，注射的DNA的一些拷贝整合到了青蛙的基因组中。这发生在.apprx中。10%-30%的病例注射环状DNA和.apprx。注射线状DNA的病例有60%-70%。通过对大肠杆菌的再转化，从注射的囊胚中回收了环状质粒。对该DNA的限制性内切酶分析表明，大多数注入的环状DNA在青蛙胚胎中复制后没有被修饰。非洲爪哇胚胎的DNA高度甲基化。注射的DNA序列即使在青蛙胚胎中经历了许多复制周期后也没有重新甲基化。从注射的DNA中可以检测到RNA转录物，直到囊胚发育的后期。
The fate of circular and linear DNA molecules following microinjection into the cytoplasm of fertilized eggs of Xenopus laevis was analyzed. Recombinant plasmids containing sea urchin histone genes (pSp 102), Drosophila ADH genes (sAC-1) and SV40 (SV2 CAT) replicate during the development of the injected frog embryo. In contrast, pBR322 either as monomers or multimers does not appear to replicate as efficiently. Injected circular DNA were not detectable by the gastrula stage of development, although there were several examples in which these molecules persisted until larval stages. In 90% of the cases, injected linear DNA persisted as discrete molecules into early embryonic stages. A portion of the DNA sequences complementary to injected linear and circular molecules was detected comigrating with the high-MW cellular frog DNA (48 kb [kilobases] or larger) from midcleavage stages onward. Restriction enzyme analysis of DNA from injected embryos suggested some copies of the injected DNA were integrated into the frog genome. This occurred in .apprx. 10%-30% of the cases of injected circular DNA and .apprx. 60%-70% of the cases of injected linear DNA. Circular plasmids were rescued from the injected blastulae by retransforming Escherichia coli. Restriction enzyme analysis of this DNA suggested that the majority of injected circular DNA were not modified following replication in the frog embryo. The DNA of Xenopus embryos was highly methylated. Injected DNA sequences were not methylated de novo even after many replication cycles in the frog embryo. RNA transcripts from the injected DNA were detectable by the late blastula stage of development.