PLASMIDS CAN STABLY TRANSFORM YEAST MITOCHONDRIA LACKING ENDOGENOUS MTDNA
PLASMIDS CAN STABLY TRANSFORM YEAST MITOCHONDRIA LACKING ENDOGENOUS MTDNA
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DOI：
10.1073/pnas.85.19.7288
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发表时间：
1988-10-01
影响因子：
11.1
通讯作者：
MCMULLIN, TW
中科院分区：
文献类型：
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作者：
FOX, TD;SANFORD, JC;MCMULLIN, TW
细菌质粒上携带的线粒体基因 oxi1 已被用于转化缺乏 mtDNA (rho0) 的酵母菌株的线粒体。通过高速微粒轰击引入质粒 DNA 后，其表现与正常酵母 rho mtDNA 的已知特性完全一致。与天然rho菌株中保留的mtDNA序列一样，转化体中的质粒DNA被重新形成多联体，其大小与野生型mtDNA没有区别。转化体中的 oxiI 序列被源自质粒的限制性位点包围，而野生型 mtDNA 中不存在这些限制性位点。这些“合成rho-”菌株中的oxiI遗传信息可以在“标记拯救”后通过与携带适当oxi1点突变的rho+ mtDNA重组而在二倍体中表达，也可以在质粒衍生的oxi1序列和删除oxi1的rho+ mtDNA异质性二倍体生长过程中反式表达。通过转化产生这种“合成rho-”菌株的能力将允许将体外产生的突变转移至野生型rho+ mtDNA，并检查反式改变基因的功能。
The mitochondrial gene oxi1, carried on a bacterial plasmid, has been used to transform the mitochondria of a yeast strain lacking mtDNA (rho0). The plasmid DNA behaved in a manner entirely consistent with the known properties of normal yeast rho- mtDNA after its introduction by high-velocity microprojectile bombardment. Like the mtDNA sequences retained in natural rho- strains, the plasmid DNA in the transformants was reiterated into concatemers whose size was indistinguishable from that of wild-type mtDNA. The oxiI sequences in the transformants were surrounded by restriction sites derived from the plasmid that were not present in wild-type mtDNA. oxiI genetic information in these "synthetic rho-" strains could be expressed in diploids either after "marker rescue" by recombination with rho+ mtDNA carrying an appropriate oxi1 point mutation or in trans during the growth of diploids heteroplasmic for both the plasmid-derived oxi1 sequences and rho+ mtDNA with oxi1 deleted. The ability to generate such "synthetic rho-" strains by transformation will allow transfer of mutations generated in vitro to wild-type rho+ mtDNA as well as examination of the function of altered genes in trans.