Hybridization kinetics between immobilized double-stranded DNA probes and targets containing embedded recognition segments

Abstract

We have investigated the time-dependent strand displacement activity of several targets with double-stranded DNA probes (dsProbes) of varying affinity. Here, the relative affinity of various dsProbes is altered through choices in hybridization length (11–15 bases) and the selective inclusion of center mismatches in the duplexes. While the dsProbes are immobilized on microspheres, the soluble, 15 base-long complementary sequence is presented either alone as a short target strand or as a recognition segment embedded within a longer target strand. Compared to the short target, strand displacement activity of the longer targets is slower, but still successful. Additionally, the longer targets exhibit modest differences in the observed displacement rates, depending on the location of recognition segment within the long target. Overall, our study demonstrates that the kinetics of strand displacement activity can be tuned through dsProbe sequence design parameters and is only modestly affected by the location of the complementary segment within a longer target strand.

A liquid-crystal-based DNA biosensor for pathogen detection

“…However, clear homeotropic domains appeared 5 min after the thermally degraded ssDNA bc was injected ( Fig. 8 aii), indicating that the high-molecular-weight target ssDNA had difficulties in hybridizing with the ssDNA probe , consistent with other reports 33 . Figure 8 aiii,iv show the POM images of TEM DTAB/DNA s functionalized with ITS1 and ITS4, respectively, after injection of the thermally unzipped ssDNA fg into the TEM DTAB/DNA cell.…”

“…Thus, ssDNA with complete mismatch (ssDNA mm ) does not change the initial planar orientation owing to difficulty in hybridization with ssDNA probe , but ssDNAs with 2- and 3-bp mismatches (DNA 2bpmm and DNA 3bpmm ) cause scattered homeotropic domains, indicating that the degree of mismatch in ssDNA target influences the LC configuration through different degrees of hybridization. The specificity for the target ssDNA is known to decrease as the chain length of the probe ssDNA decreases 33 . However, this TEM grid biosensor differentiated complete, 2-bp, and 3-bp mismatched ssDNAs from a complementary (target) ssDNA, even in 16-mer oligonucleotides, indicating that it provides high specificity.…”

Synthetic Oligonucleotide Probes for Detection of Autoimmune Antibodies

“…Finally, diverse nano-particles have been successfully applied in the development of new serotyping strategies based on synthetic unmodified dsDNA antigens [39][40][41]. For example, Baker and coworkers investigated the time-dependent strand displacement activity of several targets with dsDNA probes of varying affinity [41]. In this work the relative affinity of various probes was altered through choices in hybridization length (11-15 bases) and the selective inclusion of center mismatches in the duplexes.…”

“…Additionally, the longer targets exhibited modest differences in the observed displacement rates, depending on the location of recognition segment within the long target. Similar assays can be applied for studies of the antibody-dsDNA binding kinetics and also to determine essential probe sequence design parameters for efficient target binding [41].…”

Three‐dimensional modeling and simulation of DNA hybridization kinetics and mass transport as functions of temperature in a microfluidic channel

“…The time required for hybridization of target ssDNA to complimentary ssDNA probes immobilized on defined surfaces in microfluidic channels or microarrays is often dominated by relatively slow diffusional processes . Even with recirculating flow, hybridization times greater than 1 h are needed to reach the desired signal intensity for optical detection of hybridized, fluorescently tagged DNA .…”