Establishment of a DNA quantitation method based on DTT-crystal violet
10.3969/j.issn.1673-4130.2025.13.007
- VernacularTitle:基于DTT-结晶紫的DNA定量方法的建立
- Author:
Yuqin YE
1
;
Huang CAO
;
Xufeng WU
;
Yao TAN
;
Quanfu MA
Author Information
1. 武汉科技大学医学院,湖北武汉 430065;湖北省妇科恶性肿瘤临床医学研究中心,湖北武汉 430070
- Keywords:
dithiothreitol;
crystal violet;
DNA quantitation
- From:
International Journal of Laboratory Medicine
2025;46(13):1575-1580
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish a DNA quantitation method based on dithiothreitol(DTT)-crystal vio-let.Methods DTT was used to decolorize crystal violet,mixed with different concentrations of λ-DNA and salmon sperm DNA standard samples or concentration standard samples,and the absorbance was read at 595 nm wavelength by microplate reader,and compared with the results of ultraviolet absorbance method.DTT-crystal violet method and ultraviolet absorbance method were used to compare the concentration of plasmid samples and the concentration of genomic DNA samples of cervical exfoliated cells.The protein tolerance of the two methods was assessed by simulating protein contaminants with bovine serum albumin(BSA).Results In the quantification of λ-DNA and salmon sperm DNA,the DTT-crystal violet method had a robust linear correlation between the absorbance at 595 nm and DNA concentration(r2＞0.95),and the measured concentrations of the standard samples were not significantly different from the theoretical concentrations of the prepared standard samples(P＞0.05).There was no significant difference in the concentration of plasmid samples measured by DTT-crystal violet method and ultraviolet absorption method(P＞0.05).The concen-tration of DNA samples from cervical exfoliated cells measured by ultraviolet absorption method was positive-ly correlated with that by DTT-crystal violet method(r=0.94,P＜0.01).The concentration of the standard sample containing BSA 1 μg/μL measured by ultraviolet absorption method was higher than that of the con-trol sample,and the difference was statistically significant(P＜0.01),whereas the DTT-crystal violet method was not significantly affected(P＞0.05).Conclusion DTT-crystal violet method has obvious advantages over the existing DNA quantitation method,and is suitable for DNA quantitative analysis in scientific research and clinic.
