Real-time observation of a single DNA digestion by λ exonuclease under a fluorescence microscope field

被引：55

作者：

Matsuura, S ^{[1
]}

Komatsu, J ^{[1
]}

Hirano, K ^{[1
]}

Yasuda, H ^{[1
]}

Takashima, K ^{[1
]}

Katsura, S ^{[1
]}

Mizuno, A ^{[1
]}

机构：

[1] Toyohashi Univ Technol, Dept Ecol Engn, Aichi 4418580, Japan

来源：

NUCLEIC ACIDS RESEARCH | 2001年 / 29卷 / 16期

关键词：

D O I：

10.1093/nar/29.16.e79

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

A fluorescence microscopy technique has been developed to visualize the behavior of individual DNA and protein molecules. Real-time direct observation of a single DNA molecule can be used to investigate the dynamics of DNA-protein interactions, such as the DNA digestion reaction by lambda exonuclease. In conventional methods it is impossible to analyze the dynamics of an individual lambda exonuclease molecule on a DNA because they can only observe the average behavior of a number of exonuclease molecules. Observation of a single molecule, on the other hand, can reveal processivity and binding rate of an individual exonuclease molecule. To evaluate the dynamics of lambda exonuclease, a stained lambda DNA molecule with one biotinylated terminal was fixed on an avidin-coated coverslip and straightened using a d.c. electric field. Microscopic observation of digestion of a straightened DNA molecule by lambda exonuclease revealed that the DNA digestion rate was similar to 1000 bases/s and also demonstrated high processivity.

引用

页数：5

共 21 条

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