Micronuclear DNA from Paramecium tetraurelia: serotype 51 A gene has internally eliminated sequences.
Micronuclear DNA from Paramecium tetraurelia: serotype 51 A gene has internally eliminated sequences.
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来自四尿草履虫的微核 DNA：血清型 51 A 基因已内部消除序列。
DOI：
10.1111/j.1550-7408.1992.tb04448.x
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发表时间：
1992
期刊：
影响因子：
--
通讯作者：
PreerJr,JR
中科院分区：
文献类型：
--
作者：
Preer,LB;Hamilton,G;PreerJr,JR
A method for the isolation of micronuclear DNA fromParamecium tetraureliahas been developed. After cell lysis, a low speed centrifugation at 1,000gis used to remove all of the unbroken cells and macronuclei and approximately two thirds of the macronuclear fragments. Next a higher speed centrifugation of 9,000gsediments the micronuclei and frees them from small particulates and soluble constituents. Advantage is then taken of the fact that micronuclei have a lower density than do macronuclear fragments in 45%–60% Percoll. Micronuclei float to the top during centrifugation at 24,000g, while macronuclear fragments sediment. After several cycles of centrifugation in Percoll, the micronuclei, although heavily contaminated with cytoplasmic components, are essentially free of macronuclei and macronuclear fragments. Micronuclear DNA can then be extracted from the suspension. The whole procedure is very rapid and in about an hour micronuclear and macronuclear DNA can be separated. About 2μg of micronuclear DNA can be obtained from 6 times 107paramecia. We find that there are internal sequences in the micronuclearAgene DNA in wild type cells which are eliminated when the micronuclei develop into macronuclei. They yield unique restriction fragments for micronuclei and macronuclei. Therefore the purity of the preparations is easily monitored by probing Southern blots of restriction enzyme‐digested DNA with the clonedAgene. No differences have been found between the micronuclearAgene in wild type and the d48 mutant.
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