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Wright Lab

Mfg Tool for Analyzing DNA Secondary Structure

This website provides directions for a software tool called 'mfg', which is used to analyze DNA secondary structures during transcription.

Description of the mfg Program and Its Uses

Introduction to the mfg computer program (Download PDF)

The mfg computer program was created assuming that mutable bases have evolved to be unpaired and located in the most stable secondary structures formed during transcription (1) Mfg interfaces with the mfold program (2), which simply folds successive single-stranded segments of a specified length, while mfg temporarily halts this process when a stem is encountered, as each fold is programmed to initiate at an unpaired base. For each successive base mfg reports the Gvalue of the most stable secondary structure in which it is unpaired and the percent of total folds in which it is unpaired (i.e., percent unpaired). The Mutability Index (MI) of each unpaired base is the product of these two variables. Mfg simulates transcription in the sense that snap shot views of the most stable secondary structures within an assigned window size are identified for each unpaired base in a progressive manner, simulating RNA polymerase progression along the template strand during transcription (see Figure 1). Window size is a user-determined parameter which relates to the level of transcription and the amount of single-stranded DNA available for folding; increased rates of transcription correlate with increased mutation frequency (Figure 1).These simulations involve a running competition for shared nucleotides between successive, inter-converting secondary structures of different stabilities, and patterns emerge of repetitive stem loop-structures suggesting that certain high stability structures predominate by forming repeatedly. Presumably, the highest stability stem-loop structures exist in higher concentrations than those of lower stability, which are iterated fewer times.

RNA polymerase progression along the template strand during transcription

Uses of mfg

The mfg computer program interfaces with the mfold program (2) and has demonstrated predictive value, e.g., the correlation between mutation frequency and the rate of transcription (4, 5), and between mutation frequency and base exposure (%unpaired) (3). The program can be used to (a)predict the location of stems and loops for further analyses of predicted secondary structures; and (b) to determine the optimal window size (i.e. optimal base exposure) for further analyses of a mutable base during simulated transcription. While this tutorial explains the use (a) of mfg, further instructions are necessary for explaining the procedures involved in the other types of analyses(manuscript in preparation).

Technical note

A confusing but essential aspect of the mfg program is that it is programmed to report the most stable secondary structure in which each base exists unpaired. Thus, a paired base in a stem is treated as an unpaired base with respect to reporting the most stable secondary structure in which it is unpaired (e.g., a structure which may have been formed much earlier in the sequence). Therefore, if one selects a paired base in a stem, mfg reports a structure of no relevance to the immediate sequence segment under analysis. Modifications of the program will be made to output concerning paired bases, and a manuscript is in preparation targeted to biochemists who may wish to use mfg, but do not need to know precisely how it works.

The user-friendliness of this software is under development and will be improved. For additional information and support, contact or


1. Wright, B.E., Reschke, D.K., Schmidt, K.H., Reimers, J.M. & Knight, W. Predicting mutation frequencies in stem-loop structures of derepressed genes: implications for evolution. Mol Microbiol48, 429-41 (2003).
2. Markham, N.R. & Zuker, M. DINAMelt web server for nucleic acid melting prediction. Nucleic Acids Res 33, W577-81 (2005).
3. Wright, B.E., Reimers, J.M., Schmidt, K.H. & Reschke, D.K. Hypermutable bases in the p53 cancer gene are at vulnerable positions in DNA secondary structures. Cancer Res 62, 5641-4(2002).
4. Wright, B.E., Longacre, A. & Reimers, J.M. Hypermutation in derepressed operons of
Escherichia coli K-12. Proc Natl Acad Sci U S A 96, 5089-94 (1999).
Reimers, J.M., Schmidt, K.H., Longacre, A., Reschke, D.K. & Wright, B.E. Increased
transcription rates correlate with increased reversion rates in leuB and argH Escherichia coli auxotrophs. Microbiology 150, 1457-66 (2004).

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