DNA Thermal Denaturation Research Articles

Spectroscopic Investigation of the Quadruplex DNA‐Binding Properties of 9‐Aryl‐Substituted Isoquinolinium Derivatives

AbstractThe spectroscopic investigation of the binding properties of berberine‐type 9‐aryl‐substituted isoquinolinium derivatives with G‐quadruplex DNA (G4‐DNA) are presented. Photometric titrations show that these ligands bind with high affinity to the telomeric G4‐DNA form 22AG (K=1.0–44×105 M−2). Furthermore, fluorimetric analysis of thermal DNA denaturation (FRET melting) reveals a significant thermal stabilization of G4‐DNA 22AG upon association with the methoxy‐substituted derivatives. As an analytically useful property, the derivatives with a phenyl substituent or with additional electron‐donating groups show a very weak fluorescence intensity, which increased significantly upon G4‐DNA complexation (fluorescence light‐up effect). Additional time‐resolved fluorescence spectroscopy indicated increased fluorescence lifetimes of the DNA‐bound 9‐(4‐methoxyphenyl)‐substituted derivative, when interacting with the quadruplex‐forming strand 22AG. Notably, the changes of the steady‐state and time‐resolved emission properties of the ligand are more pronounced with G4‐DNA than with duplex DNA so that the combination of these complementary methods may be used for the selective G4‐DNA detection.

Mitochondriotropic agents conjugated with NSAIDs through metal ions against breast cancer cells

Two copper(I) polymorphs of formula [Cu(SALH)(TPP)3] (1a and 1b) were prepared by the conjugation of the Non-Steroidal Anti-Inflammatory Drug (NSAID) salicylic acid (SALH2) with the mitochondriotropic agent triphenylphosphine (TPP) via metal ion. For comparison, the isomorph [Ag(SALH)(TPP)3] (2) was prepared. The conjugates 1a, 1b and 2 were characterized by melting point (m.p.), Attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy, Ultraviolet-Visible (UV–Vis) spectroscopy and nuclear magnetic resonance (1H NMR). The crystal structures of 1a, 1b and 2 were confirmed by X-ray diffraction crystallography (XRD). The ex vivo binding affinity of 1–2 towards CT (calf thymus)-DNA was studied by UV, fluorescence, viscosity and DNA Thermal Denaturation studies. Their inhibitory activity against lipoxygenase (LOX) (an enzyme which is mainly located in the mitochondrion) was determined. The in vitro activity of 1–2 was evaluated against human breast cancer cell lines MCF-7 (hormone depended (HD)) and MDA-MB 281 (hormone independent (HI)) cells. Compounds 1–2 inhibit stronger than cisplatin the cancerous cells. The molecular mechanism of action of 1–2 was suspected by the MCF-7 cells morphology and confirmed by DNA fragmentation, Acridine Orange/Ethidium Bromide (AO/EB) Staining and mitochondrial membrane permeabilization tests.

Impact of the zinc complexation of polytopic polyaza ligands on the interaction with double and single stranded DNA/RNA and antimicrobial activity.

Metal complexes have gained a huge interest in the biomedical research in the last decade because of the access to unexplored chemical space with regards to organic molecules and to present additional functionalities to act simultaneously as diagnostic and therapeutic agents. Herein, we evaluated the interaction of two polytopic polyaza ligands and their zinc complexes with DNA and RNA by UV thermal denaturation, fluorescence and circular dichroism spectroscopic assays. The zinc coordination was investigated by X-ray diffraction and afforded the structure of the binuclear zinc complex of PYPOD. Thermal denaturation of DNA and RNA and fluorimetry analysis revealed preferential binding of the zinc-PHENPOD complexes towards GC-containing DNA in contrast to the free ligands. On the other hand, PYPOD metal complexes, compared to the free ligand, stabilized AT-based DNA (B-form) better than AU-RNA (A-form). With regards to single stranded RNA, the binuclear complex of PHENPOD and the free ligand can efficiently identify polyadenylic acid (poly A) among other RNA sequences by circular dichroism spectroscopy. The antimicrobial activity in S. aureus and E. coli bacteria showed the highest activity for the free ligands and their trinuclear zinc complexes. This work can provide valuable insights into the impact of the nuclearity of polytopic polyaza ligands in the binding to DNA/RNA and the antimicrobial effect.

Synthesis, DNA binding and biological evaluation of benzimidazole Schiff base ligands and their metal(ii) complexes.

Two novel benzimidazole ligands (E)-2-((4-(1H-benzo[d]imidazole-2-yl)phenylimino)methyl)-6-bromo-4-chlorophenol (L1) and (E)-1-((4-(1H-benzo[d]imidazole-2-yl)phenylimino)methyl)naphthalene-2-ol (L2) with their corresponding Cu(ii), Ni(ii), Pd(ii) and Zn(ii) complexes were designed and synthesized. The compounds were characterized by elemental, IR, and NMR (1H & 13C) spectral analyses. Molecular masses were determined by ESI-mass spectrometry, and the structure of ligand L1 was confirmed by single crystal X-ray diffraction analysis. Molecular docking was carried out for the theoretical investigation of DNA binding interactions. The results obtained were verified experimentally by UV/Visible absorption spectroscopy in conjunction with DNA thermal denaturation studies. It was observed that ligands (L1 and L2) and complexes (1-8) were moderate to strong DNA binders, as evident from the binding constants (K b). The value was found to be highest for complex 2 (3.27 × 105 M-1) and lowest for 5 (6.40 × 103 M-1). A cell line study revealed that breast cancer cells were less viable to the synthesized compounds compared to that of standard drugs, cisplatin and doxorubicin, at the same concentration. The compounds were also screened for in vitro antibacterial activity for which complex 2 showed a promising broad-spectrum effect against all tested strains of bacteria, almost in the proximity of the reference drug kanamycin, while the rest of the compounds displayed activity against selected strains.

Effect of TiO2 nanoparticles on the thermal stability of native DNA under UV irradiation

$TiO_{2}$ nanoparticles (NPs) are widely used in the environmental engineering, medicine, chemical and food industries due to their unique photocatalytic and biocidal properties. NPs may generate reactive oxygen species and, hence, have the toxic effect on the living cells via oxidative stress. An external UV irradiation may magnify the photocatalytic properties of $TiO_{2}$ NPs. In this regard, we have analyzed the influence of $TiO_{2}$ NPs on the conformation and thermal stability of native DNA in a buffer solution without and under UV irradiation exploiting absorption spectroscopy with DNA thermal denaturation in the range of 20-94 degrees Celsius. Upon DNA heating from about 25 to 44 degrees Celsius, we have observed the stabilization of DNA duplex in the presence of $TiO_{2}$ NPs. This additional biopolymer stabilization indicates that partial DNA unwinding appears as a result of the direct binding of the biopolymer to NPs. We showed that the performed UV treatment of DNA during 3 hours leads to partial unwinding of the biopolymer structure. The NPs injection to the biopolymer solution induced the additional effect on the DNA thermal stability under UV irradiation. The performed analysis of the experimental data suggests that the nature of the impact of NPs on the biopolymer is complex.

Effect of SDS Micelles on Actinomycin D - DNA Complexes.

DNA thermal denaturation was evaluated as a measure of the effect of antitumor drug actinomycin D on the stability of the double helix and also the effect of SDS micelles on actinomycin D - DNA complexes. The results indicated that the melting temperature of DNA was dependent on drug concentration, increasing with actinomycin D concentration. High thermal stabilization (about 10 °C) of the DNA helix after the association with actinomycin D clearly demonstrates the intercalative binding mode. The presence of SDS micelles leads to the release of intercalated actinomcyin D molecules from DNA double helix and their further relocation in surfactant micelles. These results highlighted that the drug release can be controlled in time and by varying the concentration and nature of surfactant.

Synthesis and biological evaluation of novel tetranuclear cyclopalladated complex bearing thiosemicarbazone scaffold ligand: Interactions with double‐strand DNA, coronavirus, and molecular modeling studies

AbstractThreading intercalators are a novel class of materials that carry two substituents along the diagonal positions of an aromatic ring. When bound to DNA, these substituents project out in DNA grooves. Tetranuclear complexes appear to be promising threading intercalators for developing therapeutics against cancer and viral infections that require high nucleic acid binding affinity. The objective of this work was to prepare the thiosemicarbazone scaffold ligand [4‐ClC6H4CHN=NC(S)NHPh] and tetranuclear cyclopalladated complex [Pd(4‐ClC6H4CHN=NC(S)NHPh)4] and to characterize the compounds by elemental analysis, 1D and 2D NMR, HRMS, and IR spectroscopy. The calf thymus DNA (CT‐DNA) binding properties of the compounds were investigated in vitro under simulated physiological conditions using UV–vis spectroscopy, emission spectral titration, methylene blue competitive binding, circular dichroism, DNA thermal denaturation, DNA binding, and coronavirus interactions using molecular simulation. The compounds showed cytotoxic effect against both human breast (MCF‐7) and colorectal (HCT116) cancer cells in a dose‐dependent manner. We demonstrated that the compounds are promising for DNA threading intercalation binders with large DNA binding constants on the order of 107 M−1 magnitude.

Bio-inspired nickel nanoparticles of pyrimidine-Schiff base: In vitro anticancer, BSA and DNA interactions, molecular docking and antioxidant studies

In this work, interactions of pyrimidine derivative Schiff base ligand (DMPMM) were studied and its stabilized powder nickel nanoparticles (DMPMM-NiNPs) were synthesized and various biological studies were evaluated. DNA binding studies of CT-DNA with prepared compounds in Tris-HCl/NaCl buffer were carried out by traditional UV-Visible and fluorescence spectroscopic methods, viscosity measurements and cyclic voltammetry. Results showed that the small scale of DMPMM had less activity to interact with biological systems and when it assembled on nickel nanoparticles surface the activity increased. Thermal denaturation and sonochemical denaturation studies of DNA with the presence and the absence of our compounds also were done by UV-Visible spectroscopic method and its results indicated that the synthesized compounds increased the denaturation temperature. BSA binding studies of synthesized compounds were done by UV-Visible and fluorescence spectroscopy. Molecular docking of prepared ligand and its nanoparticles with biomolecules (DNA and BSA) were studied. Antimicrobial studies of the DMPMM and DMPMM-NiNPs were carried out by Agar-Agar well diffusion method. Anticancer studies results evidenced that the synthesized DMPMM-NiNPs had good selectivity to control the growth of cancer cells without damaging the normal cells. Various antioxidant scavenging studies results have shown that DMPMM and DMPMM-NiNPs have significant antioxidant activity. Highlights Stable and solid nickel nanoparticles were prepared. The size of the prepared nickel nanoparticles was nearly 3 to 8 nm. Organic ligand capped nickel nanoparticles interacted with DNA and BSA. Ni nanoparticles increased the denaturation temperature of DNA. It was found to have good anticancer activity with fewer side effects than cisplatin. Communicated by Ramaswamy H. Sarma

Synthesis of 10-O-aryl-substituted berberine derivatives by Chan-Evans-Lam coupling and investigation of their DNA-binding properties.

Eleven novel 10-O-aryl-substituted berberrubine and berberine derivatives were synthesized by the Cu2+-catalyzed Chan–Evans–Lam coupling of berberrubine with arylboronic acids and subsequent 9-O-methylation. The reaction is likely introduced by the Cu2+-induced demethylation of berberrubine and subsequent arylation of the resulting 10-oxyanion functionality. Thus, this synthetic route represents the first successful Cu-mediated coupling reaction of berberine substrates. The DNA-binding properties of the 10-O-arylberberine derivatives with duplex and quadruplex DNA were studied by thermal DNA denaturation experiments, spectrometric titrations as well as CD and LD spectroscopy. Fluorimetric DNA melting analysis with different types of quadruplex DNA revealed a moderate stabilization of the telomeric quadruplex-forming oligonucleotide sequence G3(TTAG3)3. The derivatives showed a moderate affinity towards quadruplex DNA (Kb = 5–9 × 105 M−1) and ct DNA (Kb = 3–5 × 104 M−1) and exhibited a fluorescence light-up effect upon complexation to both DNA forms, with slightly higher intensity in the presence of the quadruplex DNA. Furthermore, the CD- and LD-spectroscopic studies revealed that the title compounds intercalate into ct DNA and bind to G4-DNA by terminal stacking.

Berberrubine Phosphate: A Selective Fluorescent Probe for Quadruplex DNA.

A phosphate-substituted, zwitterionic berberine derivative was synthesized and its binding properties with duplex DNA and G4-DNA were studied using photometric, fluorimetric and polarimetric titrations and thermal DNA denaturation experiments. The ligand binds with high affinity toward both DNA forms (Kb = 2–7 × 105 M−1) and induces a slight stabilization of G4-DNA toward thermally induced unfolding, mostly pronounced for the telomeric quadruplex 22AG. The ligand likely binds by aggregation and intercalation with ct DNA and by terminal stacking with G4-DNA. Thus, this compound represents one of the rare examples of phosphate-substituted DNA binders. In an aqueous solution, the title compound has a very weak fluorescence intensity (Φfl < 0.01) that increases significantly upon binding to G4-DNA (Φfl = 0.01). In contrast, the association with duplex DNA was not accompanied by such a strong fluorescence light-up effect (Φfl < 0.01). These different fluorimetric responses upon binding to particular DNA forms are proposed to be caused by the different binding modes and may be used for the selective fluorimetric detection of G4-DNA.

Rational Design and Synthesis of Naphthalene Diimide Linked Bis-Naphthalimides as DNA Interactive Agents.

A molecular modeling assisted rational design and synthesis of naphthalene diimide linked bis-naphthalimides as potential DNA interactive agents is described. Chemical templates incorporating naphthalene diimide as a linker in bis-naphthalimide motif were subjected to molecular docking analysis at specific intercalation and telomeric DNA G-quadruplex sites. Excellent results were obtained, which were better than the standards. A short and convenient synthetic route was employed to access these hybrids experimentally, followed by evaluation of their ability to cause thermal denaturation of DNA and cytotoxic properties along with ADME predictions. The obtained results provided useful insights and two potential molecules were identified for further development.

DNA thermal denaturation by polymer field theory approach: effects of the environment

We analyse the effects of the environment (solvent quality, presence of extended structures - crowded environment) that may have impact on the order of the transition between denaturated and bounded DNA states and lead to changes in the scaling laws that govern conformational properties of DNA strands. We find that the effects studied significantly influence the strength of the first order transition. To this end, we re-consider the Poland-Scheraga model and apply a polymer field theory to calculate entropic exponents associated with the denaturated loop distribution. For the d = 3 case, the corresponding diverging ε = 4-d expansions are evaluated by restoring their convergence via the resummation technique. For the space dimension d = 2, the exponents are deduced from mapping the polymer model onto a two-dimensional random lattice, i.e., in the presence of quantum gravity. We also show that the first order transition is further strengthened by the presence of extended impenetrable regions in a solvent that restrict the number of the macromolecule configurations.

Binding Characteristics of Dibenzo[a,h]Anthracene with DNA In Vitro: Investigated by Spectroscopic and Magnetic Bead Methods

In this study, the interacting characteristics of dibenzo[a,h]anthracene (DBA) with free DNA in vitro were investigated through multiple conventional spectroscopic techniques. UV–vis absorption spectroscopy was used to initially explore whether there was an interaction between DBA and DNA. Fluorescence spectroscopy was exploited to explore the binding characteristics and binding force between DBA and DNA. Circular dichroism (CD) spectra and DNA thermal denaturation measurement are utilized to further explore the binding mode between DBA and DNA. Magnetic bead separation technique coupled with fluorescence microscopic observation visually demonstrated the binding between DBA and DNA, and the residual amount of DBA was measured by fluorescence intensity. The results show that DBA can bind to DNA in vitro through intercalation mode. In the process of forming DBA–DNA complex, hydrogen bond and van der Waals force play a major role. The binding constant (K A) and binding sites (n) are 2.039 × 105 L/mol and 1.0451 at 25 °C, respectively. Moreover, DNA can be used to capture DBA in the solution based on DBA–DNA intercalation, and its removal efficiency is 98.24%. This work can provide new insights for the establishment of a novel polycyclic aromatic hydrocarbons (PAHs) elimination method based on PAHs–DNA intercalation.

Synthesis and investigation of quadruplex-DNA-binding, 9-O-substituted berberine derivatives.

A small series of five novel berberine derivatives was synthesized by the Cu-catalyzed click reaction of 9-propargyladenine with 9-O-(azidoalkyl)berberine derivatives. The association of the resulting berberine–adenine conjugates with representative quadruplex-forming oligonucleotides 22AG dA(G3TTA)3G3 and a2 d(ACAG4TGTG4)2 was examined with photometric and fluorimetric titrations, thermal DNA denaturation analysis, and CD spectroscopy. The results from the spectrometric titrations indicated the formation of 2:1 or 1:1 complexes (ligand:G4-DNA) with log Kb values of 10–11 (2:1) and 5–6 (1:1), which are typical for berberine derivatives. Notably, a clear relationship between the binding affinity of the ligands with the length of the alkyl linker chain, n, was not observed. However, depending on the structure, the ligands exhibited different effects when bound to the G4-DNA, such as fluorescent light-up effects and formation of ICD bands, which are mostly pronounced with a linker length of n = 4 (with a2) and n = 5 (with 22AG), thus indicating that each ligand–G4-DNA complex has a specific structure with respect to relative alignment and conformational flexibility of the ligand in the binding site. It was shown exemplarily with one representative ligand from the series that such berberine–adenine conjugates exhibit a selective binding, specifically a selectivity to quadruplex DNA in competition with duplex DNA, and a preferential thermal stabilization of the G4-DNA forms 22AG and KRAS. Notably, the experimental data do not provide evidence for a significant effect of the adenine unit on the binding affinity of the ligands, for example, by additional association with the loops, presumably because the adenine residue is sterically shielded by the neighboring triazole unit.

DNA and BSA binding study of an optically pure rosin derivative and its two copper(II) complexes

To explain the differences found in cytotoxicity, interactions of an optically pure rosin derivative 2-amino-5-dehydroabietyl-1,3,4-thiadiazole (L) and its two copper(II) complexes [CuL4Cl]Cl·2CH2Cl2·H2O (1) and [CuL4Br]Br·2CH2Cl2 (2) with CT DNA and bovine serum albumins (BSA) have been studied. Emission spectral (K SV = 1.80 × 103 L mol−1 (L), 2.43 × 105 L mol−1 (1) and 2.31 × 105 L mol−1 (2)), DNA thermal denaturation and viscosity measurements reveal that 1 and 2 bind DNA through intercalation with 1 exhibiting a slight higher DNA binding ability, while L bound with DNA through groove interaction. Fluorescence and CD spectral data indicate that 1 and 2 have higher BSA affinity than L. Molecular docking analysis was carried out to explore the interactions of L, 1 and 2 toward DNA and BSA. All the findings reveal that the DNA and BSA binding affinity followed the order of 1 > 2 > L. In addition, their antitumor abilities were in this order, too.

Pyrimidine Derivative Schiff Base Ligand Stabilized Copper and Nickel Nanoparticles by Two Step Phase Transfer Method; in Vitro Anticancer, Antioxidant, Anti-Microbial and DNA Interactions.

Pyrimidine derivative Schiff base ligand (DPMC) stabilized metal nanoparticles of copper (DPMC-CuNPs) and nickel (DPMC-NiNPs) were synthesized by modified Brust-Schiffrin technique, which is a two-step phase transfer assisted synthesis. The prepared metal nanoparticles were confirmed by UV-Visible and Infrared spectroscopy. The size, surface morphology and the quality of the DPMC and its MNPs were analyzed by Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM) methods respectively. Electrochemical behavior of the DPMC-CuNPs and DPMC-NiNPs was analyzed by cyclic voltammetry method. DNA binding studies of the synthesized compounds with CT-DNA were examined by four different techniques such as UV-Visible and emission spectroscopy, cyclic voltametry and viscometric measurments. Thermal denaturation and sono-chemical denaturation studies of DNA with the DPMC, DPMC-CuNPs and DPMC-NiNPs results also suggest the synthesized compounds have good DNA binding ability. Various antioxidant scavenging studies results shows that DPMC and its copper and nickel nanoparticles have significant antioxidant activity. Antimicrobial studies of the DPMC and its MNPs were studied by Agar-Agar well diffusion method. Anticancer studies of the DPMC and its MNPs show that the DPMC-CuNPs and DPMC-NiNPs have significant anticancer activity with least toxicity than the standard drug cis-platin. Graphical Abstract.

(E)-(1-(4-Ethoxycarbonylphenyl)-5-(3,4-dimethoxyphenyl)-3-(3,4-dimethoxystyryl)-2-pyrazoline: Synthesis, Characterization, DNA-Interaction, and Evaluation of Activity Against Drug-Resistant Cell Lines

(E)-1-(4-Ethoxycarbonylphenyl)-5-(3,4-dimethoxyphenyl)-3-(3,4-dimethoxystyryl)-2-pyrazoline was synthesized via the cyclization reaction between the monocarbonyl curcuminoid (2E,6E)-2,6-bis(3,4-dimethoxybenzylidene)acetone and ethyl hydrazinobenzoate in high yield and purity (&gt;95% by High-performance liquid chromatography (HPLC)). The compound has been fully characterized by 1H, 13C NMR, FTIR, UV-Vis and HRMS and its activity was evaluated in terms of its potential interaction with DNA as well as its cytotoxicity against resistant and non-resistant tumor cells. Both DNA thermal denaturation and DNA viscosity measurements revealed that a significant intercalation binding takes place upon treatment of the DNA with the synthesized pyrazoline, causing an increase in melting temperature by 3.53 ± 0.11 °C and considerable DNA lengthening and viscosity increase. However, neither re-sensitisation of Doxorubicin (DO X)-resistant breast cancer and multidrug resistance (MDR) reversal nor synergistic activity with DOX by potentially increasing the DOX cell killing ability was observed.

Influence of Glass and Air on Our Perception of DNA

The chemical and physical properties of glass do not allow it to be considered as an inert material. Therefore, it is necessary to discuss the ability of laboratory glassware to influence the state and properties of aqueous solutions, including DNA solutions. As a demonstration of this ability, it is shown here how contact with glass affects the state of salts in aqueous solutions. In terms of the topic under discussion, it is especially important that sodium salts are very sensitive to these contacts. Thus, it is shown here that contact with glass can affect the formation of fibers from sodium salts of DNA, which were mainly used in X-ray studies aimed at determining the molecular structure and parameters of DNA molecules. Special attention is paid to the fact that laboratory glassware is also sensitive to contact with aqueous solutions, including DNA solutions. That such sensitivity may be important to biologists is also shown here. In particular, it was shown how ignoring the sensitivity of silica glass, from which the photometric cells are made, to electrization gave rise to misconceptions about the spectral properties of aqueous solutions, including DNA solutions. The effect of air dissolved in aqueous DNA solutions on the spectral properties of both these solutions and the photometric cells containing them is also shown. In particular, it was shown that UV absorption of aqueous DNA solutions, which does not contain air, is completely insensitive to heating, and UV absorption of aqueous DNA solutions, which are saturated with air, is supersensitive to heating. In other words, it is shown here that our understanding of the thermal denaturation of DNA, which is reflected in the UV absorption of its aqueous solutions, was formed due to these two types of sensitivity. In the end, a conclusion is proposed here about the significant contribution of glass and air to the creation of traditional ideas about the structure and properties of DNA.

Microscopic description of DNA thermal denaturation

We propose a microscopic model describing the process of DNA thermal denaturation. The process consists of the splitting of DNA base pairs, or nucleotides, resulting in the separation of two complementary DNA strands. In contrast to the previous modelling attempts we take into account the states of all base pairs of DNA which in fact imposes the microscopic nature of the approach. The model is a linear integro-differential non-autonomous equation describing the dynamics of probability density which characterizes the distances between the bases within individual base pairs. The non autonomous structure of the equation comes from the dependence on temperature that may be in the general case a (given) function of time. To our knowledge it is the first model taking into account not only the strength of double and triple hydrogen bonds between the complementary bases but also the stacking interactions between neighborhood base pairs. In the present paper the basic preliminary properties of the model including the existence, uniqueness and stability results in various cases are discussed. Moreover the symmetrized version of the model is considered and the “macroscopic limit” is studied. The performed numerical simulations reproduce the sigmoid shape of DNA melting curves and reveal the appearance of experimentally observed denaturation bubbles.

Thermomechanical responses of microfluidic cantilever capture DNA melting and properties of DNA premelting states using picoliters of DNA solution

Melting or thermal denaturation of a DNA molecule and the different bubble-rich, premelting DNA states that serve as a precursor for DNA thermal denaturation are vital events in DNA thermophysics. In this study, we employ cantilever-based sensing to firstly pinpoint the occurrence of DNA melting and identify the temperature Tm characterizing the melting. Very importantly, this sensing is carried out with an extremely small volume (∼picoliters) of DNA sample with the cantilever demonstrating an extremely high sensitivity on the order of mJ/g⋅K corresponding to pico-Joules of energy input. Secondly, this same large sensitivity of the cantilever is used to quantify the hitherto unknown thermophysical properties of the bubble-rich DNA premelting states. In fact, for both the melting and premelting states, the cantilever provides a framework to calculate the specific heat capacity and the storage and loss moduli of the cantilever-DNA-solution system, thereby establishing a platform for quantifying DNAs' thermo-mechanical behavior.