Generation of Multiblock Copolymers by PCR: Synthesis, Visualization and Nanomechanical Properties (original) (raw)
Related papers
Recent progress on DNA block copolymer
Chinese Chemical Letters, 2017
Organic polymers are combined with DNA resulting DNA block copolymers (DBCs) that can simultaneously show the properties of the polymer and DNA. We will discuss some examples of recent developments in the syntheses, structure manipulations, and applications of DBCs.
Three-Dimensional Organization of Block Copolymers on “DNA-Minimal” Scaffolds
Journal of the …, 2012
Here, we introduce a 3D-DNA construction method that assembles a minimum number of DNA strands in quantitative yield, to give a scaffold with a large number of single-stranded arms. This DNA frame is used as a core structure to organize other functional materials in 3D as the shell. We use the ring-opening metathesis polymerization (ROMP) to generate block copolymers that are covalently attached to DNA strands. Site-specific hybridization of these DNA-polymer chains on the single-stranded arms of the 3D-DNA scaffold gives efficient access to DNA-block copolymer cages. These biohybrid cages possess polymer chains that are programmably positioned in three dimensions on a DNA core and display increased nuclease resistance as compared to unfunctionalized DNA cages.
2010
Introduction of nucleic acids into cells is an important biotechnology research field which also holds great promises for therapeutic applications. One of the key steps in the gene delivery process is compaction of DNA into nanometric particles. The study of DNA condensing properties of three linear cationic triblock copolymers poly(ethylenimine-b-propylene glycol-b-ethylenimine), namely , indicates that proper DNA condensation is driven both by the charge and the size of the respective cationic hydrophilic linear polyethylenimine (LPEI) and neutral hydrophobic polypropylene glycol (PPG) parts. Atomic Force Microscopy was used to investigate the interactions of the triblock copolymers with plasmid DNA at the single molecule level and to enlighten the mechanism involved in DNA condensation.
Stimuli-responsive organization of block copolymers on DNA nanotubes
Chemical Science, 2012
I. General 3-hydroxypicolinic acid, acetic acid, ammonium citrate, acetic acid, boric acid, cyanogen bromide (5M in acetonitrile), EDTA, formamide, urea, 4-morpholineethanesulfonic acid (MES), magnesium chloride, StainsAll ® , and tris(hydroxymethyl)-aminomethane (Tris) were purchased from Aldrich. Nucleoside (dA, T, dC, dG) derivatized 1000 Å and 2000Å LCAA-CPG supports with loading densities between 25-40 µmol/g, 5-ethylthiotetrazole and reagents used for automated DNA synthesis, Sephadex G-25 (super fine, DNA grade) 3'-alkyne-modifier and 5'-amino-modifier C6 were purchased from Glenn Research. 1000Ǻ Phosphate-CPG was purchased from ChemGenes. Exonuclease VII (ExoVII, source: recombinant) was used as purchased from BioLynx Incorporated. 40% acrylamide/bis-acrylamide 19:1 solution and agarose were purchased from BioShop. A RepliPHI TM Phi29 reagent set was purchased from Epicentre Biotechnologies for rolling circle amplification. Grade SPI-1 highly ordered pyrolytic graphite (HOPG) was purchased from SPI Supplies INC. and Ruby Red mica sheets (1 x 3") were purchased from B & M MICA CO., while etched silicon cantilevers (OMCL-AC160TS) were purchased from Olympus for AFM imaging. 1 x TBE buffer is composed of 90 mM Tris, 90 mM boric acid and 11 mM EDTA with a pH ~8.3. 1 x TA Mg buffer is composed of 40 mM Tris and 7.6 mM MgCl 2 •6H 2 O. The pH was adjusted to 8.0 using glacial acetic acid. 1 x MES Mg buffer is composed of 250 mM MES, 20 mM MgCl 2 , with pH 7.6. II. Instrumentation Standard automated oligonucleotide solid-phase synthesis was performed on a BioAutomation MerMade MM6 DNA synthesizer. UV-Vis measurements were performed with a BioTek Synergy HT microplate reader. Gel electrophoresis experiments were carried out on an acrylamide 20 x 20 cm vertical Hoefer 600 electrophoresis unit. Thermals anneal cycles and enzymatic digestions were conducted using a Flexigene Techne 96 well thermocycler. AFM was performed with a MultiMode TM SPM connected to a Nanoscope TM IIIa controller, from the Digital Instruments Veeco Metrology Group. Circular dichroism (CD) spectra were recorded on a JASCO J-810 spectrophotometer. Dynamic light scattering (DLS) measurements were performed on a Brookhaven Instruments Corporation system equipped with a
Biomaterials, 2005
A series of comb-type copolymers comprised of various polycation backbones and dextran (Dex) side chains were prepared to study the DNA/copolymer interaction. While the cationic copolymers with a lower degree of dextran grafts maintained an ability to condense DNA molecules into a globule form those with a higher degree of dextran grafting interacted with DNA without inducing DNA condensation. The structural differences in cationic backbones diversely influenced DNA hybridization as evaluated by circular dichroism (CD) spectrometry and UV-melting analyses. The copolymer having a polyallylamine (PAA) backbone induced B-A-type transformation of DNA duplex, whereas the copolymers having either a-poly(l-lysine) (aPLL) or e-poly(l-lysine) (ePLL) backbone induced B-C-type transformation. The PAA copolymer is the first example of the artificial polymer that induces B-A-type transformation under physiologically relevant condition. UV-melting analyses of DNA strands indicated that the aPLL copolymers showed the highest stabilization efficacy toward poly(dA) Á poly(dT) duplex and poly(dA) Á 2poly(dT) triplex without affecting reversibility of inter DNA association. Melting temperatures (T m ) of the triplex increased from 38 C to 99 C by the addition of the aPLL copolymer with an appropriate grafting degree. While the PAA copolymers had higher density of cationic groups along the backbone than aPLL copolymers, these copolymers moderately increased T m of the DNA triplex. The PAA copolymer caused considerable hysteresis in thermal melting/reassociation processes. Note that the ePLL copolymers increased T m of the DNA triplex and not the duplex, suggesting their potential as a triplex selective stabilizer. Chemical structures of the cationic backbones of the copolymers were characteristically affected on the copolymer/DNA interaction even if their backbones were surrounded by abundant side chains (> 65 wt%) of dextran. The study suggested that tailor-made design of ''functional polycounterion'' is a strategy to engineer molecular assembling of DNA. r
DNA Sequence and Length Dictate the Assembly of Nucleic Acid Block Copolymers
The self-assembly of block copolymers is often rationalized by structure and microphase separation; pathways that diverge from this parameter space may provide new mechanisms of polymer self-assembly. Here, we show that the sequence and length of single-stranded DNA directly influence the self-assembly of sequence-defined DNA block copolymers. While increasing the length of DNA led to predictable changes in self-assembly, changing only the sequence of DNA produced three distinct structures: spherical micelles (spherical nucleic acids, SNAs) from flexible poly(thymine) DNA, fibers from semi-rigid mixed-sequence DNA, and networked superstructures from rigid poly(adenine) DNA. The secondary structure of poly(adenine) DNA strands drives a temperature-dependent polymerization and assembly mechanism: copolymers stored in an SNA reservoir form fibers after thermal activation, which then aggregate upon cooling to form interwoven networks. DNA is often used as a programming code that aids in...
Polymeric effects on DNA condensation by cationic polymers observed by atomic force microscopy
Colloids and Surfaces B: Biointerfaces, 2010
Compaction of DNA by condensing agents can provide insights into DNA assembly processes, which is keenly related to the essence of gene transfection and gene therapy in vivo. In this paper, the morphology of different cationic polymer/DNA complexes was studied by using atomic force microscopy (AFM), which is keen to the mechanism of DNA condensation induced by amine-based cationic block copolymers with poly(poly(ethylene glycol) methyl ether methacrylate). It is found that the structures and dimensions of condensing agent/DNA complexes are sensitively dependent on the condensing agents. The size of DNA aggregates can be affected appreciably by polymers rather than monomers. The amount of nitrogen elements per polymer unit, rather than the molecular weights of polymers, appears to be more effective on the dimension of the condensates. The impact of the copolymer chain structures on the DNA aggregates indicates an effective venue for regulating the dimensions and structures of the DNA condensates, which is beneficial for optimizing delivery systems for gene transfection.
DNA-nanoparticle assemblies go organic: Macroscopic polymeric materials with nanosized features
Journal of Nanobiotechnology, 2012
Background: One of the goals in the field of structural DNA nanotechnology is the use of DNA to build up 2-and 3-D nanostructures. The research in this field is motivated by the remarkable structural features of DNA as well as by its unique and reversible recognition properties. Nucleic acids can be used alone as the skeleton of a broad range of periodic nanopatterns and nanoobjects and in addition, DNA can serve as a linker or template to form DNA-hybrid structures with other materials. This approach can be used for the development of new detection strategies as well as nanoelectronic structures and devices. Method: Here we present a new method for the generation of unprecedented all-organic conjugated-polymer nanoparticle networks guided by DNA, based on a hierarchical self-assembly process. First, microphase separation of amphiphilic block copolymers induced the formation of spherical nanoobjects. As a second ordering concept, DNA base pairing has been employed for the controlled spatial definition of the conjugated-polymer particles within the bulk material. These networks offer the flexibility and the diversity of soft polymeric materials. Thus, simple chemical methodologies could be applied in order to tune the network's electrical, optical and mechanical properties.
We herein report the synthesis and characterization of ABC-type triblock copolymers containing two complementary association motifs and investigate their folding into well-defined polymeric nanoparticles under diluted conditions via intramolecular orthogonal hydrogen bonding. The precursor ABC-type triblock copolymers are prepared via reversible addition–fragmentation chain transfer (RAFT) polymerization bearing primary alkyl bromide on A, protected alkyne on B, and protected hydroxyl pendant groups on the C units. The dithioester groups of the RAFT polymers are quantitatively removed by radical-induced reduction before the side-chain functionalization. The complementary motifs, i.e., Hamilton wedge (HW, A block), benzene-1,3,5-tricarboxamide (BTA, B block), and cyanuric acid (CA, C block), are incorporated into the linear triblock copolymers side chains via postfunctionalization. The self-assembly processes of the HW and CA supramolecular motifs are followed by nuclear magnetic resonance (1H NMR) spectroscopy at ambient and elevated temperature in various solvents. The helical BTA stack formation is monitored by circular dichroism (CD) spectroscopy. In addition, the final aggregates formed by these two orthogonal forces, namely HW-CA pseudo-cross-linking and BTA stacking, are characterized by static and dynamic light scattering (SLS and DLS) as well as atomic force microscopy (AFM).