Automatic analysis of 2D polyacrylamide gels in the diagnosis of DNA polymorphisms

Robert Koprowski,Karolina Chwiałkowska,Mirosław Kwaśniewski,Anna Korzyńska,Zygmunt Wróbel

doi:10.1186/1475-925x-12-68

Abstract

IntroductionThe analysis of polyacrylamide gels is currently carried out manually or automatically. In the automatic method, there are limitations related to the acceptable degree of distortion of lane and band continuity. The available software cannot deal satisfactorily with this type of situations. Therefore, the paper presents an original image analysis method devoid of the aforementioned drawbacks.MaterialThis paper examines polyacrylamide gel images from Li-Cor DNA Sequencer 4300S resulting from the use of the electrophoretic separation of DNA fragments. The acquired images have a resolution dependent on the length of the analysed DNA fragments and typically it is MG×NG=3806×1027 pixels. The images are saved in TIFF format with a grayscale resolution of 16 bits/pixel. The presented image analysis method was performed on gel images resulting from the analysis of DNA methylome profiling in plants exposed to drought stress, carried out with the MSAP (Methylation Sensitive Amplification Polymorphism) technique.ResultsThe results of DNA polymorphism analysis were obtained in less than one second for the Intel Core™ 2 Quad CPU Q9300@2.5GHz, 8GB RAM. In comparison with other known methods, specificity was 0.95, sensitivity = 0.94 and AUC (Area Under Curve) = 0.98.ConclusionsIt is possible to carry out this method of DNA polymorphism analysis on distorted images of polyacrylamide gels. The method is fully automatic and does not require any operator intervention. Compared with other methods, it produces the best results and the resulting image is easy to interpret. The presented method of measurement is used in the practical analysis of polyacrylamide gels in the Department of Genetics at the University of Silesia in Katowice, Poland.

Highlights

The analysis of polyacrylamide gels is currently carried out manually or automatically
Material: This paper examines polyacrylamide gel images from Li-Cor DNA Sequencer 4300S resulting from the use of the electrophoretic separation of DNA fragments
The presented image analysis method was performed on gel images resulting from the analysis of DNA methylome profiling in plants exposed to drought stress, carried out with the MSAP (Methylation Sensitive Amplification Polymorphism) technique

Summary

Conclusions

The proposed methodology for the analysis and processing of polyacrylamide gel images enables to perform an automatic and repeatable measurement of the position of lanes and bands. This method is superior to the previously presented methods, described in [5] and [11], in cases of gel deformation. Proper identification of bands and lanes in the described method enables to obtain the output image LDNA. Correction must be made in the image LDNA This method suggests a distinct approach to the analysis of lanes that involves the comparison of adjacent lanes directly in the image LO, which requires separate studies.

Introduction

Method