Gel Alignment and Band Scoring for DNA Fingerprinting Using Adobe® Photoshop®

Understanding the genetic relatedness among bacterial strains has a variety of practical applications, including the identification of possible sources of food-borne disease outbreaks and tracking the dissemination of specific bacterial genotypes. The genetic relatedness of bacterial strains is usually identified using molecular DNA fingerprinting techniques such as pulse-field gel electrophoresis, repetitive sequence -based PCR, and ERIC-PCR analysis (1,3,4). A key step common to all of these methods is the banding pattern analysis of DNA bands in electrophoresis gels. By interpreting these banding patterns, genetic relatedness “dendograms” or “trees” are created that allow for the identification of genetically related strains (2). Commercial software programs are available that will facilitate almost all of the steps involved in DNA banding pattern analysis, including image capture, digitization, image analysis, and interpretation. Some of these programs are sold either as stand-alone programs or as parts of the electrophoresis or image capture equipment. The prices can vary between $2000 and $15 000. Very often, because of cost considerations, many laboratories (especially those at academic institutions that have only modest operating budgets) are forced to rely on the visual examination and interpretation of the banding patterns to identify genetic relatedness among bacterial strains. The differences in the running conditions between gels on different days and by different individuals who may use different electrophoresis equipment and power supplies can create differences in the migration patterns of the DNA size markers (ladders) between different gels. This prevents an accurate gel-to-gel comparison, thereby compromising the quality of the data analysis and interpretation. We rationalized that if commonly available and relatively inexpensive software such as Adobe® Photoshop® (version 5.5 or higher) is used for the standardization and alignment of DNA banding patterns obtained in different gels, then it will help reduce human errors when aligning the DNA banding patterns on different gels. There are two critical steps for comparing DNA banding patterns, (i) the alignment (standardization) of the bands across multiple gels and (ii) the “scoring” (presence/absence) of specific bands across the different gels. The approach presented here will permit the alignment of DNA bands across multiple gels and create a grid that will facilitate the scoring of the various bands. Gel running conditions such as run times, voltage, and staining need to be rigorously standardized before electrophoresis. We recommend the use of premade gels and the loading of DNA size markers on the two outermost lanes and a center lane to help in gel alignment. This method can be used to compare as many gels as possible; however, factors such as fatigue and eye strain can play a role. Given below is a standardized protocol for the alignment of banding patterns and the creation of a grid that will facilitate the scoring of the banding patterns.