Experimental approach for development of a model system with specific information clouded by noise.

<p>Specific information of interest (orange) sandwiched by redundant information (green) is gradually clouded by vector DNA (turquoise) and cellular DNA (blue). Vector DNA and cellular DNA represent potential noise sources of increasing complexity.</p

Southern blot for 40 nt sequences.

<p>The complementary sequence is labelled with a marker. This marker enables detection of the hybridized sequence through X-ray radiography. Lanes 1, 2 and 3, unpurified MEA 40 nt product in three different amounts (20, 10, 5 uL), lane 4 and 5, the ssDNA product from modified isothermal amplification method with a single primer. Lane 6–8, purified MEA products. Lanes 9–10, reference ssDNA sequences of 40 nt length, of oligonucleotide composition as the intended 40 nt sequence.</p

Modified end amplification (MEA) technique.

<p>The DNA template (1) requires a well defined end point of the template (2) that can be produced for instance by means of a restriction enzyme (red). Denaturation (3) at 95°C and annealing (4) at 50°C are similar to conventional PCR. Taq polymerase (blue) generates a complementary ssDNA oligonucleotide fragment from a single primer (red) during the extension step (5). The complementary ssDNA oligonucleotide fragment is extended until the end of the source strand (6). Use of a single primer results in linear amplification (7). There are n copies of complementary ssDNA oligonucleotide fragments after n cycles.</p

Specific molecular recognition.

<p>The figure illustrates the sequence specific temperature dependent nucleic acid hybridization of a DNA. The strand (TGACATGCTAATC) is complementary to ssDNA strand (ACTGTCGATTAG).</p

Application of modified end amplification (MEA) approach, identification, retrieval and verification of specific information.

<p>Oligonucleotide sequences are extracted with the MEA technique and purified using standard nucleic acid purification and extraction procedures. They are immobilized on a southern blot membrane by hybridizing to complementary oligonucleotide sequences with a marker. The presence of small fragments of defined length from MEA technique results in highly specific hybridization, and noise is minimized.</p

Hamming Distance as a Concept in DNA Molecular Recognition

DNA microarrays constitute an in vitro example system of a highly crowded molecular recognition environment. Although they are widely applied in many biological applications, some of the basic mechanisms of the hybridization processes of DNA remain poorly understood. On a microarray, cross-hybridization arises from similarities of sequences that may introduce errors during the transmission of information. Experimentally, we determine an appropriate distance, called minimum Hamming distance, in which the sequences of a set differ. By applying an algorithm based on a graph-theoretical method, we find large orthogonal sets of sequences that are sufficiently different not to exhibit any cross-hybridization. To create such a set, we first derive an analytical solution for the number of sequences that include at least four guanines in a row for a given sequence length and eliminate them from the list of candidate sequences. We experimentally confirm the orthogonality of the largest possible set with a size of 23 for the length of 7. We anticipate our work to be a starting point toward the study of signal propagation in highly competitive environments, besides its obvious application in DNA high throughput experiments

Symbols correspond to insertion bases (A red crosses; C green circles; G blue stars; T cyan triangles)

The mean profile (black line), obtained from the moving average (including all 4 insertion types) over positions - 2 to + 2 shows the common positional dependence. Insertions to the left and to the right of an identical base (bulges – see text) result in identical probe sequences. and Deviation profiles. Positional influence is mostly eliminated by subtraction of the mean profile. Elevated intensities are observed for bulges (e.g. C insertions at positions 11 to 15, 6 to 7 and 18 to 20 or G insertions at positions 4 to 5 and 7 to 8). A very distinct increase of the hybridization signal is observed for C insertions into the subsequence TCCCCT in the middle of the sequence. As shown in bulges (red markers) have significantly higher intensities compared to bulges (blue markers).<p><b>Copyright information:</b></p><p>Taken from "Impact of point-mutations on the hybridization affinity of surface-bound DNA/DNA and RNA/DNA oligonucleotide-duplexes: Comparison of single base mismatches and base bulges"</p><p>http://www.biomedcentral.com/1472-6750/8/48</p><p>BMC Biotechnology 2008;8():48-48.</p><p>Published online 13 May 2008</p><p>PMCID:PMC2435543.</p><p></p

Impact of point-mutations on the hybridization affinity of surface-bound DNA/DNA and RNA/DNA oligonucleotide-duplexes: Comparison of single base mismatches and base bulges-9

O the target oligonucleotide COM. Markers depict the substituent base type (A red crosses; C green circles; G blue stars; T cyan triangles). The black line indicates the 'mean profile' (moving average of all mismatch hybridization signals over positions - 2 to + 2). PM probes, included as control to detect erroneous bias, have the largest hybridization signals (at a level of about 0.38 a.u.). The variation of the PM probe intensities also provides an estimate for the error of the measurement. Errors between distant microarray features, due to gradient effects, are expected to be larger than errors between the compactly arranged features corresponding to a particular defect position. Deviation profile. The strong position dependent component of the hybridization signal is eliminated by subtraction of the mean profile. Comparison of mean mismatch hybridization signals (average of the three mismatch hybridization signals at a particular defect position) at the sites of C·G base pairs to mean MM hybridization signals at the site of adjacent A·T base pairs. A marker (red star: A·T; blue circle C·G) is set in the upper row if the hybridization signal of the mismatches at the corresponding site is higher than at the adjacent site; otherwise a marker is set in the lower row. We noticed that mismatches substituting a C·G base pair usually have systematically lower hybridization signals than mismatches substituting a neighboring A·T base pair.<p><b>Copyright information:</b></p><p>Taken from "Impact of point-mutations on the hybridization affinity of surface-bound DNA/DNA and RNA/DNA oligonucleotide-duplexes: Comparison of single base mismatches and base bulges"</p><p>http://www.biomedcentral.com/1472-6750/8/48</p><p>BMC Biotechnology 2008;8():48-48.</p><p>Published online 13 May 2008</p><p>PMCID:PMC2435543.</p><p></p

Boxes indicate the interquartile range (from the 25th to 75th percentile) containing 50% of the data

Whiskers extend to a maximum value of 1.5 times the interquartile range from the boxes ends. differ significantly with a 95 percent confidence. Data processing: raw intensity data, solution-background correction, subtraction of the mean profile, normalization of the defect-type dependent deviations from the mean profile by division by the standard deviation of the defect profile (see Methods section). The mismatch types with the lowest hybridization signals are those (T·G, C·C, T·C, A·C, G·G) where C·G base pairs are affected by the mismatch defect. The only exception is A·G. The positive tails of this and other distributions seem to originate from stabilizing C·G base pairs next to the defect. (standard deviation assuming that the various MM nearest-neighbor types are equally distributed).<p><b>Copyright information:</b></p><p>Taken from "Impact of point-mutations on the hybridization affinity of surface-bound DNA/DNA and RNA/DNA oligonucleotide-duplexes: Comparison of single base mismatches and base bulges"</p><p>http://www.biomedcentral.com/1472-6750/8/48</p><p>BMC Biotechnology 2008;8():48-48.</p><p>Published online 13 May 2008</p><p>PMCID:PMC2435543.</p><p></p

(a) Ranking order of DNA/DNA mismatch binding affinities (extracted from Fig

4). (b) As anticipated the ranking order for DNA/DNA MMs obtained from the smaller subset of probe sequences (Fig. 9A) is very similar. The ranking order for the analogue RNA/DNA MM duplex stabilities (c) (extracted from Fig. 9B) reveals significant differences in comparison to (b). In part (d) MM-types are ordered according to the hybridization signal differences between RNA/DNA and DNA/DNA MMs (as extracted from Additional file ). Purine-purine MMs (purine bases highlighted in blue) display the largest decrease of binding afinities with respect to other MM-types.<p><b>Copyright information:</b></p><p>Taken from "Impact of point-mutations on the hybridization affinity of surface-bound DNA/DNA and RNA/DNA oligonucleotide-duplexes: Comparison of single base mismatches and base bulges"</p><p>http://www.biomedcentral.com/1472-6750/8/48</p><p>BMC Biotechnology 2008;8():48-48.</p><p>Published online 13 May 2008</p><p>PMCID:PMC2435543.</p><p></p