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Competitive binding-based optical DNA mapping for fast identification of bacteria--multi-ligand transfer matrix theory and experimental applications on Escherichia coli.

Nilsson AN, Emilsson G, Nyberg LK, Noble C, Stadler LS, Fritzsche J, Moore ER, Tegenfeldt JO, Ambjörnsson T, Westerlund F - Nucleic Acids Res. (2014)

Bottom Line: Our identification protocol introduces two theoretical constructs: a P-value for a best experiment-theory match and an information score threshold.The developed methods provide a novel optical mapping toolbox for identification of bacterial species and strains.The protocol does not require cultivation of bacteria or DNA amplification, which allows for ultra-fast identification of bacterial pathogens.

View Article: PubMed Central - PubMed

Affiliation: Department of Astronomy and Theoretical Physics, Lund University, Sölvegatan 14A, 223 62 Lund, Sweden.

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(A) Experimental raw kymograph for T4 DNA at 1:150 YOYO:netropsin in 0.05× TBE. Fluorescent images of DNA molecules were recorded at different times (time along the vertical axis). The sample was mixed in 5× TBE and diluted. (B) Aligned kymograph. (C) DNA barcode consisting of 20 lines generated from the average of the experimental kymograph. (D) Comparing the experimental (black) and theoretical (gray) barcodes.
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Figure 4: (A) Experimental raw kymograph for T4 DNA at 1:150 YOYO:netropsin in 0.05× TBE. Fluorescent images of DNA molecules were recorded at different times (time along the vertical axis). The sample was mixed in 5× TBE and diluted. (B) Aligned kymograph. (C) DNA barcode consisting of 20 lines generated from the average of the experimental kymograph. (D) Comparing the experimental (black) and theoretical (gray) barcodes.

Mentions: To obtain a time-averaged experimental ‘barcode’, one must account for center-of-mass diffusion in the channel and conformational fluctuations. To that end, we applied a slightly modified version of the ‘local box stretching’ approach in (30). We complemented the algorithm by applying a moving average on the experimental signal, for alignment purposes. We also introduced a rough method for aligning the start and finish pixel of the region containing the DNA (see Supplementary Information for details and for an analysis of the noise properties of the aligned experimental barcodes). See Figure 4 for an example of the result of the alignment and data fitting.


Competitive binding-based optical DNA mapping for fast identification of bacteria--multi-ligand transfer matrix theory and experimental applications on Escherichia coli.

Nilsson AN, Emilsson G, Nyberg LK, Noble C, Stadler LS, Fritzsche J, Moore ER, Tegenfeldt JO, Ambjörnsson T, Westerlund F - Nucleic Acids Res. (2014)

(A) Experimental raw kymograph for T4 DNA at 1:150 YOYO:netropsin in 0.05× TBE. Fluorescent images of DNA molecules were recorded at different times (time along the vertical axis). The sample was mixed in 5× TBE and diluted. (B) Aligned kymograph. (C) DNA barcode consisting of 20 lines generated from the average of the experimental kymograph. (D) Comparing the experimental (black) and theoretical (gray) barcodes.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4150756&req=5

Figure 4: (A) Experimental raw kymograph for T4 DNA at 1:150 YOYO:netropsin in 0.05× TBE. Fluorescent images of DNA molecules were recorded at different times (time along the vertical axis). The sample was mixed in 5× TBE and diluted. (B) Aligned kymograph. (C) DNA barcode consisting of 20 lines generated from the average of the experimental kymograph. (D) Comparing the experimental (black) and theoretical (gray) barcodes.
Mentions: To obtain a time-averaged experimental ‘barcode’, one must account for center-of-mass diffusion in the channel and conformational fluctuations. To that end, we applied a slightly modified version of the ‘local box stretching’ approach in (30). We complemented the algorithm by applying a moving average on the experimental signal, for alignment purposes. We also introduced a rough method for aligning the start and finish pixel of the region containing the DNA (see Supplementary Information for details and for an analysis of the noise properties of the aligned experimental barcodes). See Figure 4 for an example of the result of the alignment and data fitting.

Bottom Line: Our identification protocol introduces two theoretical constructs: a P-value for a best experiment-theory match and an information score threshold.The developed methods provide a novel optical mapping toolbox for identification of bacterial species and strains.The protocol does not require cultivation of bacteria or DNA amplification, which allows for ultra-fast identification of bacterial pathogens.

View Article: PubMed Central - PubMed

Affiliation: Department of Astronomy and Theoretical Physics, Lund University, Sölvegatan 14A, 223 62 Lund, Sweden.

Show MeSH
Related in: MedlinePlus