Base-pairing mismatches

Examine DNA. How many base pairs does it contain Starting from one end, write down the sequence of bases in one strand. Write down the sequence in the complementary strand. Is this a proper DNA fragment, or does it contain base-pair mismatches ... 

Fig. 5 Charge transfer occurs through the DNA /r-stack and is strongly dependent on minor base stack perturbations. Photoinduced electron transfer is observed from tethered intercalated ethidium to a rhodium intercalator bound to DNA up to 30 A away. The efficiency of electron transfer is drastically reduced in the presence of a stacking disruption, here a single base-pair mismatch... 

Modulation of DNA structure and dynamics is also possible using base-pair mismatches. Mismatches exert little influence on the global structure of B-DNA duplexes. Locally, the extent of base stacking perturbation depends sensitively on the nature of the mismatch [139-141]. Therefore, while a CA mismatch introduces a significant distortion in local stacking, the well-stacked GA mismatch is, by many criteria, barely perceptible. The dynamics of mismatched base-pairs may also be significantly distinct from matched Watson-Crick base pairs [9]. We exploit these features of DNA mismatches to probe the sensitivity of DNA-mediated CT to base structure and dynamics. 

The innate sensitivity of DNA-mediated CT to perturbations in the TT-stack has prompted us to employ this chemistry as a probe of stacking structure and dynamics. We have developed a new class of DNA-based diagnostic tools that diagnose DNA mutations such as single base-pair mismatches and lesions, analyze DNA-protein interactions, and probe the sequence-depen-dent dynamics and flexibility of DNA. These applications rely on electrochemical probing of CT in DNA films self-assembled on gold electrodes. 

The dynamics of intercalation of small molecules with DNA, groove binding and binding to specific sites, such as base pair mismatches have been studied by stopped-flow,23,80 108 temperature jump experiments,26,27,94 109 120 surface plasmon resonance,121 129 NMR,86,130 135 flash photolysis,136 138 and fluorescence correlation spectroscopy.64 The application of the various techniques to study the binding dynamics of small molecules will be analyzed for specific examples of each type of binding. 

Fig. 14.7 (a) Measured resonant mode spectral shift for increasing concentrations of target oligonucleotides. The line fit is a Michaelis Menton curve with a Kd of 2.9 nM and a maximum resonant wavelength shift of 5.1 pm. (b) Measured resonant mode shift for oligonucleotides of increasing number of base pair mismatches with the capture probe. Reprinted from Ref. 32 with permission. 2008 Elsevier... 

Watson-Crick wobble base pair mismatch mismatch mismatch... 

The use of nanoparticles to form bridges between two electrodes was also utilized to detect DNA hybridization, including single base-pair mismatches, by Mirkin and... 

Shen, P. Huang, H.V. (1993). Effect of base-pair mismatches on recombination via the RecBCD pathway. Molec. Gen. Genetics 218, 358-360. 

Fig. 2. (A) Analysis of human adult skeletal (HAS) and cardiac muscle (HAC) transcripts by an oligonucleotide array representing all 363 exons of the human titin gene. Left Comparison of results obtained with HAS (top) and HAC (bottom) transcripts reveals large blocks of exons that are only positive in skeletal muscle. Right Examples of constitutively expressed exons (exon 5 and 7), cardiac specific exons (11 and 49), and skeletal muscle specific exons (156 and 210). 5MM a 50mer from exon 5 including five base-pair mismatches as a control for hybridization...

Melt temperature measurements indicated that the thermodynamic stability of the probe molecules can be adjusted using the oligomer matrix to achieve lower Tm values by as much as 5 °C, with retention of selectivity for discrimination of single base pair mismatches [14]. The enhanced temperature sensitivity may ultimately provide for the development of analytical devices of extremely high selectivity. Devices containing immobilized... 

Thermal denaturation profiles were done using one of the molecular beacon structures in solution, without the alkyl chain linker attached to the 3 -thymidine residue. The fully complementary and single base pair mismatched sequence targets were investigated. The differences of Tm from the melt profiles showed that single base pair mismatches could not be discriminated from the complementary target material. Melt curves were also... 

Linear oligonucleotide probes have been immobilized along the length of fibres as well as on the distal end of fibres. Healey et al. [88] described the immobilization of three different oligonucleotide probes in acrylamide polymer matrices at the distal ends of optical fibres. Base pair mismatches could be distinguished on the same sensor after a 20-min hybridization with a detection limit of 0.2 nM [88]. More recently, zeptomole ( 600 molecule) detection limits have been obtained by using etched distal ends of optical fi-... 

Cotton R, Rodrigues NR, Campbell RD. Reactivity of cytosine and thymine in single-base-pair mismatches with hydroxylamine and osmium tetroxide and its application to the study of mutations. Proc Natl Acad Sci USA 1988 85 4397-4401. 

Kisko JL, Barton JK. Recognition of DNA base pair mismatches by a cyclomethylated Rh(III) intercalator. Inorg Chem 2000 39 4942-9. 

Junicke H, Hart JR, Kisko J, Glebov O, Kirsch IR, Barton JK. A rhodium(III) complex for high-affinity DNA base-pair mismatch recognition. Proc Natl Acad Sci USA 2003 100 3737 12. 

An estimate of the extent of base-pair mismatch between hybridized DNA strands can be made by measuring the thermal stability of the reassociated duplexes. Place hybridized filters in fresh incubation buffer and heat at 5° increments to 90°. At each 5° increase, remove a small sample of the buffer and count. Controls are reassociated homologous DNAs. The amount of nucleotide mispairing has been estimated at 1% for each degree the Tm of the heterologous duplex is lowered (Arm).2 31... 

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