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DNA and Aging -with special reference to mitochondria

DNA 和衰老——特别是线粒体

基本信息

批准号：
04836011
负责人：
ASANO Kimiko
金额：
$ 1.28万
依托单位：
KYOTO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1992
资助国家：
日本
起止时间：
1992 至 1993
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04836011/
关键词：
mitochondria DNA LINE limited-diet ad libitum heart cerebrum cerebellum kidney ヌクレアーゼ

项目摘要

Aging-dependent changes of mitochondrial DNA and nuclear DNA (mtDNA/nDNA) ratios were measured using several tissues of rats. For accurate determination, we used LINE (long highly-repetitive sequence) which is present in nuclear DNA of rats. A 2 kbp LINE fragment which was obtained by Sall and PstI digestion of purified DNA was used for biotinylated probe (probe N) synthesis. mtDNA fragment A (6.2 kbp) of EcoRI digestion was used as a mitochondria-specific probe. Using these two probes, mtDNA/nDNA ratio can be obtained by a single experiment. The ratio of each point was calculated as average of at least three independent measurements.In heart muscle, mtDNA/nDNA ratio of 30 month-old rats decreased to about 30% of young (2 month) samples as in the case of rat liver. Experimental results described above and pervious years were those from rats on 60%-limited diet. In case of those fed ad libitum, age-dependent decrease of mtDNA/nDNA ratio was the same as limited samples in liver, however, slower decrease of the ration in heart muscle was observed. In kidney, the ratio stayd at the same level in case of the limited diet, but some decrease was evident with those fed ad libitum. In case of rats which were fed ad libitum, decrease of mtDNA was observed in cerebrum, but not in cerebellum.

用大鼠不同组织测定线粒体DNA和核DNA （mtDNA/nDNA）比值随年龄变化的变化。为了准确测定，我们使用了存在于大鼠核DNA中的LINE（长高度重复序列）。纯化DNA经small和PstI酶切得到2 kbp的LINE片段，用于生物素化探针（探针N）的合成。EcoRI酶切的mtDNA片段A （6.2 kbp）作为线粒体特异性探针。使用这两种探针，可以通过一次实验获得mtDNA/nDNA比值。每个点的比率计算为至少三次独立测量的平均值。在心肌中，30月龄大鼠的mtDNA/nDNA比率下降到年轻（2个月）样品的30%左右，与大鼠肝脏的情况相同。上述和前几年的实验结果是60%限制饮食的大鼠。在自由喂养的情况下，肝脏mtDNA/nDNA比值的年龄依赖性下降与有限样本相同，而心肌mtDNA/nDNA比值的下降速度较慢。在肾脏方面，限饲时该比率保持在相同水平，而自由饲喂时该比率有所下降。随意喂养大鼠时，大鼠脑区mtDNA减少，小脑区mtDNA未见减少。