DNA Renaturation Research Articles

Profiling a single-stranded DNA region within an rDNA segment that affects the loading of bacterial condensin.

Bacterial condensin preferentially loads onto single-stranded DNA (ssDNA) invitro and onto rDNA invivo to support proper chromosome compaction. Thus, the actively transcribing rDNA would provide the ssDNA region for the loading of bacterial condensin. We attempted to detect the ssDNA region in the rrnI gene in situ. Non-denaturing sodium bisulfite treatment catalyzed the conversion of cytosines to thymines (CT-conversion) at the melted DNA of a genome. Using next-generation sequencing, we generated an average of 11,000 reads covering each cytosine on the rDNA segment. In principle, the CT-conversion rate is an accurate guide to detect ssDNA segment. We detected multiple ssDNA segments throughout the rDNA. The deletion mutations of the rDNA hindered the ssDNA formation at the 100-500bp segment downstream of the promoter. These data support the idea that the ssDNA segment plays a crucial role as the condensin-loading site and suggest the mechanism of condensin loading onto rDNA.

Glaesserella australis sp. nov., isolated from the lungs of pigs

Twenty-nine isolates of an unknown haemophilic organism were isolated from the lungs of pigs from 14 farms in Australia. Phylogenetic analyses based on the 16S rRNA gene, recN and rpoA showed a monophyletic group that was most closely related to Glaesserella parasuis and [Actinobacillus] indolicus. Whole genome sequence analysis indicated that the Glaesserella parasuis and this group, using the type strain HS4635T for comparison, showed a similarity of 30.9 % DNA-DNA renaturation. The isolates were Gram-stain-negative, NAD-dependent, CAMP-negative and were oxidase-positive, catalase-negative and produced indole but not urease. The isolates could be separated from all currently recognized haemophilic and non-haemophilic members of the family Pastuerellaceae. Key phenotypic properties were the production of indole, the lack of urease activity, production of β-galactosidase but not α-fucosidase, acid formation from (-)-d-arabinose, (+)-d-galactose, maltose and trehalose and a failure to produce acid from (-)-d-mannitol. Taken together, these data indicate that the isolates belong to a novel species for which the name Glaesserella australis sp. nov. is proposed. The type strain is HS4635T (=CCUG 71931T and LMG 30645T).

Label-free and Non-destruction Determination of Single- and Double-Strand DNA based on Quantum Weak Measurement

The process of unwinding and renaturation of DNA has been widely used in studies of nucleotide sequence organization. Compared with traditional methods for DNA unwinding and renaturation, the label-free and non-destruction detection technology is significant and desiderated. We realized an optical system based on optical rotation via weak measurement for detection of single- and double-strand state of DNA. The optical rotation, which was induced by the status change of single and double DNA strands, was exploited to modulate the preselected polarization of a weak measurement system. With this modulation, the optical rotation caused by the separation of DNA strands can be determined through the center wavelength shift of the output spectrum. By monitoring the wavelength shift in real time, the separation processes of the DNAs with different base ratio (25% and 70%) and length (4nt and 40nt), and DNAs with three terminally modified cholesterol molecules were experimentally explored in varied pH and temperature conditions. In addition, the detection limit of the DNA concentration was obtained to be 5 × 10−6 mol/L. Our work based on optical rotation detection of single- and double-strand DNA exhibits the unique advantages of real-time monitoring, label-free, non-destruction and simplicity.

Rodentibacter gen. nov. including Rodentibacter pneumotropicus comb. nov., Rodentibacter heylii sp. nov., Rodentibacter myodis sp. nov., Rodentibacter ratti sp. nov., Rodentibacter heidelbergensis sp. nov., Rodentibacter trehalosifermentans sp. nov., Rodentibacter rarus sp. nov., Rodentibacter mrazii and two genomospecies.

Rodentibacter gen. nov. is proposed based on isolation and phenotypic characterization of strains, predominantly from rodents. The strains showed 86 % or higher rpoB gene sequence similarity and indicated a genus-level relationship within Pasteurellaceae. The strains compared at 16S rRNA gene sequence level showed 93.8 % or higher similarity, and their genus-level relationship within Pasteurellaceae was confirmed by phenotypic analysis. The type species Rodentibacter pneumotropicus comb. nov. is reclassified from [Pasteurella] pneumotropica with type strain NCTC 8141T (=CCUG 12398T). Whole genomic comparison allowed the estimation of DNA-DNA renaturation. Rodentibacter heylii sp. nov. was proposed for a group that included the biovar Heyl of [Pasteurella] pneumotropica with the type strain ATCC 12555T (=CCUG 998T). A group was proposed as Rodentibacter ratti sp. nov., which included the taxon 22 of Bisgaard; the type strain is F75T (=CCUG 69665T=DSM 103977T). Taxon 41 of Bisgaard was proposed as Rodentibacter myodis sp. nov. with type strain Ac151T (=CCUG 69666T=DSM 103994T). Rodentibacter heidelbergensis sp. nov. included the type strain 1996025094T (=Ac69T) (=CCUG 69667T=DSM 103978T). A group strains of was proposed as Rodentibacter trehalosifermentans sp. nov. with type strain H1987082031T (=CCUG 69668T=DSM 104075T). Two strains including the reference strain of taxon 17 of Bisgaard that showed 16S rRNA gene similarity of 97.3 % were proposed as Rodentibacter rarus sp. nov. 2325/79T (=CCUG 17206T=DSM 103980T). Rodentibacter mrazii sp. nov. was proposed with type strain Ppn418T (Bisgaard taxon 21) (=CCUG 69669T=DSM 103979T). The eight species could be separated based on phenotypic characteristics such as NAD requirement, ornithine decarboxylase and indole formation, α-glucosidase, β-galactosidase and in acid formation from (+)-l-arabinose, (-)-d-ribose, (+)-d-xylose, myo-inositol, (-)-d-mannitol, lactose, melibiose and trehalose. Forty-six strains including taxon 48 of Bisgaard formed a monophyletic group by rpoB and 16S rRNA gene sequence analysis, but could not be separated phenotypically from R. pneumotropicus and R. heylii, and it was left as an unnamed genomospecies 1 of Rodentibacter with reference strain Ppn416. Another taxon that included 13 strains, mainly isolated from Apodemus sylvaticus, could not be separated phenotypically from R. pneumotropicus or R. heylii and was designated as genomospecies 2. Strain Ppn85 with 95 % or less rpoB gene sequence similarity and with 16S rRNA gene sequence similarity of 97 % or less to the other members of Rodentibacter was left as an unnamed singleton.

Nanocomposite DNA hydrogels with temperature sensitivity

DNA hydrogels are promising soft materials because of the distinctive properties of DNA such as biocompatibility, coil-globule transition, molecular recognition, and selective binding. However, such hydrogels synthesized so far suffer from poor mechanical properties and low dimensional stability, which limit their stress-bearing technological applications. Here, we introduce a novel concept for the preparation of high-strength double-stranded (ds) DNA hydrogels with temperature sensitivity. The hydrogels were synthesized by free-radical polymerization of N-isopropylacrylamide in aqueous Laponite dispersions containing ds-DNA in a highly entangled state. The nanoparticles of synthetic Laponite clay act as a dynamic cross-linker to form a three-dimensional physical network due to strong clay-polymer interactions. We observed that ds-DNA maintains its native structure and function during the formation of nanocomposite hydrogels. Cyclic heating/cooling experiments conducted between below and above the melting temperature of ds-DNA reveal a drastic increase in the elastic modulus of nanocomposite hydrogels due to the thermally induced denaturation and renaturation of DNA within the physical network. The hydrogels prepared at various Laponite contents display the characteristics of both poly(N-isopropylacrylamide) and ds-DNA e.g., temperature sensitivity, denaturation and renaturation of DNA strands, and they exhibit a high tensile fracture stress (up to 53 kPa) and elongation to break (up to 544%).

Theory on the Mechanism of DNA Renaturation: Stochastic Nucleation and Zipping.

Renaturation of the complementary single strands of DNA is one of the important processes that requires better understanding in the view of molecular biology and biological physics. Here we develop a stochastic dynamical model on the DNA renaturation. According to our model there are at least three steps in the renaturation process viz. nonspecific-contact formation, correct-contact formation and nucleation, and zipping. Most of the earlier two-state models combined nucleation with nonspecific-contact formation step. In our model we suggest that it is considerably meaningful when we combine the nucleation with the zipping since nucleation is the initial step of zipping and nucleated and zipping molecules are indistinguishable. Nonspecific contact formation step is a pure three-dimensional diffusion controlled collision process. Whereas nucleation involves several rounds of one-dimensional slithering and internal displacement dynamics of one single strand of DNA on the other complementary strand in the process of searching for the correct-contact and then initiate nucleation. Upon nucleation, the stochastic zipping follows to generate a fully renatured double stranded DNA. It seems that the square-root dependency of the overall renaturation rate constant on the length of reacting single strands originates mainly from the geometric constraints in the diffusion controlled nonspecific-contact formation step. Further the inverse scaling of the renaturation rate on the viscosity of reaction medium also originates from nonspecific contact formation step. On the other hand the inverse scaling of the renaturation rate with the sequence complexity originates from the stochastic zipping which involves several rounds of crossing over the free-energy barrier at microscopic levels. When the sequence of renaturing single strands of DNA is repetitive with less complexity then the cooperative effects will not be noticeable since the parallel zipping will be a dominant enhancing factor. However for DNA strands with high sequence complexity and length one needs to consider the underlying cooperative effects both at microscopic and macroscopic levels to explain various scaling behaviours of the overall renaturation rate.

Repetitive sequences: the hidden diversity of heterochromatin in prochilodontid fish.

The structure and organization of repetitive elements in fish genomes are still relatively poorly understood, although most of these elements are believed to be located in heterochromatic regions. Repetitive elements are considered essential in evolutionary processes as hotspots for mutations and chromosomal rearrangements, among other functions – thus providing new genomic alternatives and regulatory sites for gene expression. The present study sought to characterize repetitive DNA sequences in the genomes of Semaprochilodus insignis (Jardine & Schomburgk, 1841) and Semaprochilodus taeniurus (Valenciennes, 1817) and identify regions of conserved syntenic blocks in this genome fraction of three species of Prochilodontidae (Semaprochilodus insignis, Semaprochilodus taeniurus, and Prochilodus lineatus (Valenciennes, 1836) by cross-FISH using Cot-1 DNA (renaturation kinetics) probes. We found that the repetitive fractions of the genomes of Semaprochilodus insignis and Semaprochilodus taeniurus have significant amounts of conserved syntenic blocks in hybridization sites, but with low degrees of similarity between them and the genome of Prochilodus lineatus, especially in relation to B chromosomes. The cloning and sequencing of the repetitive genomic elements of Semaprochilodus insignis and Semaprochilodus taeniurus using Cot-1 DNA identified 48 fragments that displayed high similarity with repetitive sequences deposited in public DNA databases and classified as microsatellites, transposons, and retrotransposons. The repetitive fractions of the Semaprochilodus insignis and Semaprochilodus taeniurus genomes exhibited high degrees of conserved syntenic blocks in terms of both the structures and locations of hybridization sites, but a low degree of similarity with the syntenic blocks of the Prochilodus lineatus genome. Future comparative analyses of other prochilodontidae species will be needed to advance our understanding of the organization and evolution of the genomes in this group of fish.

Characterization of denaturation and renaturation of DNA for DNA hybridization

ObjectivesThe present study was designed to systematically characterize the denaturation and the renaturation of double stranded DNA (dsDNA), which is suitable for DNA hybridization.MethodsA series of physical and chemical denaturation methods were implemented on well-defined 86-bp dsDNA fragment. The degree of each denaturation was measured and the most suitable denaturation method was determined. DNA renaturation tendency was also investigated for the suggested denaturation method.ResultsHeating, beads mill, and sonication bath did not show any denaturation for 30 minutes. However probe sonication fully denatured DNA in 5 minutes. 1 mol/L sodium hydroxide (alkaline treatment) and 60% dimethyl sulfoxide (DMSO) treatment fully denatured DNA in 2-5 minutes.ConclusionsAmong all the physical methods applied, the direct probe sonication was the most effective way to denature the DNA fragments. Among chemical methods, 60% DMSO was the most adequate denaturation method since it does not cause full renaturation during DNA hybridization.

Denaturation and renaturation behaviors of short DNA in a confined space.

A deep understanding to the denaturation and renaturation behaviors of DNA in a confined state is fundamentally important to control the self-assembly of DNA in a chamber or channel for various applications. In this report, we study the denaturation and renaturation behaviors of short DNA confined in cylindrical and spherical spaces with the 3-Site-Per-Nucleotide coarse-grained DNA model applying the replica exchange molecular dynamics technology. It is found that as the confinement size decreases, the melting temperature Tm increases and the transition becomes broad. The analysis of the potential of mean force shows that the confinement increases the relative free energy of the denatured state of DNA and decreases the renaturation energy barrier. Besides the denatured and native states, the metastable parallel-stranded structure is also found. The simulation results show that the shapes of the confinement spaces and the short DNA sequences remarkably affect the renaturation behavior. In the cylindrical space, the DNA renaturation changes from random-binding to slithering-binding with the size of the confinement space decreasing. In contrast, the DNA renaturation in the spherical and symmetrical confinement space proceeds through strand binding and rolling. The relationship between the melting temperature and the confinement size, ΔTm/Tm ∼ Rc (-υ), is estimated and the exponential index υ equals about 1.32 and 1.75 in the cylindrical and spherical confinements, respectively. It is further compared with the theoretical result of the rigid rod model and a qualitative agreement with the simulation is achieved.

DNA interaction with zinc(II) ions

We focused on interactions of Zn(II) with DNA in this study. These interactions were monitored using UV/vis spectrophotometry and gel electrophoresis. Firstly, we isolated and amplified 498bp fragment of DNA. Samples were obtained by incubation of DNA fragment with Zn(II) for 60min at 25°C. After incubation, the samples were dialyzed and analyzed immediately. In this way, DNA was converted into a metal bound DNA (Zn–DNA). Interaction of Zn(II) with DNA caused change in the absorption spectrum (190–350nm) and decrease in the melting temperature (Tm) of Zn–DNA. Spectrophotometric (UV/vis) analysis showed that increasing concentrations of zinc(II) ions led to the increase in the absorbance at 200nm and decrease in absorbance at 251nm. Application of zinc(II) ions at 5.5μM concentration caused decrease in Tm for app. 7.5°C in average in comparison with control (75.5±3°C). The lowest melting temperature (60.5±2.5°C) was observed after application of zinc(II) ions at 33μM concentration. Gel electrophoresis proved significance of Zn(II) in the renaturation of DNA. Samples of Zn–DNA (15μM DNA+5.5–55μM Zn(II)) caused significant changes in the renaturation of DNA in comparison with the control, untreated DNA (15μM DNA).

Cot-based sampling of genomes for polymorphic low-copy DNA

DNA polymorphisms are powerful tools for many evolutionary and genomic studies in plants including molecular breeding. Single nucleotide polymorphisms (SNPs) are the most elemental DNA marker for genomic studies, but even with advances in DNA sequencing technology, SNP discovery remains costly and computationally demanding, especially in large genomes that are rich in repetitive DNA such as those of many plants. Here we report a method using DNA renaturation kinetics (Cot techniques), sequencing, and BLAST-based screening to identify low-copy, non-coding DNA sequences that were subsequently found to be relatively rich in polymorphisms. A total of of 63 such fragments isolated from a diploid D genome cotton species (Gossypium raimondii) revealed a higher frequency of polymorphisms than that observed for cotton expressed sequence tags or hypomethylated (PstI-susceptible) genomic DNA. While microsatellite-derived loci show still higher polymorphism rates, they often fall in repetitive elements and their sequence analysis is often complicated by alignment difficulties. The potential applications of Cot-filtered noncoding (CFNC) DNA in development of DNA markers are discussed.

Cyclic Denaturation and Renaturation of Double-Stranded DNA by Redox-State Switching of DNA Intercalators

Hybridization of complementary nucleic acid strands is fundamental to nearly all molecular bioanalytical methods ranging from polymerase chain reaction and DNA biosensors to next generation sequencing. For nucleic acid amplification methods, controlled DNA denaturation and renaturation is particularly essential and achieved by cycling elevated temperatures. Although this is by far the most used technique, the management of rapid temperature changes requires bulky instrumentation and intense power supply. These factors so far precluded the development of true point-of-care tests for molecular diagnostics. To overcome this limitation we explored the possibility of using electrochemical means to control reversible DNA hybridization by using the electroactive intercalator daunomycin (DM). We show that redox-state switching of DM altered its properties from DNA binding to nonbinding, under otherwise constant conditions, and thus altered the thermodynamic stability of duplex DNA. The operational principle was demonstrated using complementary synthetic 20mer and 40mer DNA oligonucleotides. Absorbance-based melting curve analysis revealed significantly higher melting temperatures for DNA in the presence of oxidized compared to chemically reduced DM. This difference was exploited to drive cyclic electrochemically controlled denaturation and renaturation. Analysis with in situ UV-vis and circular dichroism spectroelectrochemistry, as two independent techniques, indicated that up to 80% of the DNA was reversibly hybridized. This remarkable demonstration of electrochemical control of five cycles of DNA denaturation and renaturation, under otherwise constant conditions, could have wide-ranging implications for the future development of miniaturized analytical systems for molecular diagnostics and beyond.

Saccharomyces cerevisiae Dmc1 and Rad51 Proteins Preferentially Function with Tid1 and Rad54 Proteins, Respectively, to Promote DNA Strand Invasion during Genetic Recombination

The Saccharomyces cerevisiae Dmc1 and Tid1 proteins are required for the pairing of homologous chromosomes during meiotic recombination. This pairing is the precursor to the formation of crossovers between homologs, an event that is necessary for the accurate segregation of chromosomes. Failure to form crossovers can have serious consequences and may lead to chromosomal imbalance. Dmc1, a meiosis-specific paralog of Rad51, mediates the pairing of homologous chromosomes. Tid1, a Rad54 paralog, although not meiosis-specific, interacts with Dmc1 and promotes crossover formation between homologs. In this study, we show that purified Dmc1 and Tid1 interact physically and functionally. Dmc1 forms stable nucleoprotein filaments that can mediate DNA strand invasion. Tid1 stimulates Dmc1-mediated formation of joint molecules. Under conditions optimal for Dmc1 reactions, Rad51 is specifically stimulated by Rad54, establishing that Dmc1-Tid1 and Rad51-Rad54 function as specific pairs. Physical interaction studies show that specificity in function is not dictated by direct interactions between the proteins. Our data are consistent with the hypothesis that Rad51-Rad54 function together to promote intersister DNA strand exchange, whereas Dmc1-Tid1 tilt the bias toward interhomolog DNA strand exchange.

Electrochemistry of Nucleic Acids

Publisher Summary The electroactivity of nucleic acids was discovered several years ago. It was shown that at mercury electrodes, adenine and cytosine were reduced in ssDNA while guanine produced an anodic signal due to the oxidation of guanine reduction product. The DNA signals at mercury electrodes are highly sensitive to changes in DNA structure due to DNA denaturation and renaturation, as well as to minor structural changes resulting from DNA premelting and DNA damage. The changes in DNA structure are reflected not only by the DNA faradaic responses but also by non-faradaic signals due to adsorption/desorption of DNA. Several interesting principles have been used in the development of the DNA sensors—such as amplified electrochemical analysis, investigation of charge transport, and tracing of changes in conformation of DNA. The s ensors for DNA hybridization and DNA damage lead the field of the electrochemistry of nucleic acids. The chapter briefly summarizes some properties of the elimination voltammetry with linear scan (EVLS) method. Oscillographic polarography, at controlled alternating current, was used in the electrochemical analysis of nucleic acids.

Mathematical analysis of the real time array PCR (RTA PCR) process

Real time array PCR (RTA PCR) is a recently developed biochemical technique that measures amplification curves (like with quantitative real time Polymerase Chain Reaction (qRT PCR)) of a multitude of different templates in a sample.It combines two different methods in order to profit from the advantages of both, namely qRT PCR (real time quantitative detection) with microarrays (high multiplex capability). This enables the quantitative detection of many more target sequences than that can be done by qRT PCR. Thereby, the concentration of many different target molecules originally present in a sample can be measured. Labelled primers are used that are first elongated to form labelled amplicons in the bulk and these can hybridize to capture probes immobilized on the surface of the microarray. During each PCR cycle, there is a time window available during which the formed labelled amplicons can hybridize to the target sequences (capture probes) on the microarray surface. By detection of the fluorescence of the spots on the microarray, amplification curves comparable to qRT PCR are obtained, which can be used to deduce the information needed on the presence and the amount of targets initially present in the sample.We present a mathematical model that provides fundamental insights in the different steps of real time array PCR (RTA PCR). These findings can be used to optimize the different biochemical processes taking place.At the microarray surface specific molecules are captured and taken away from the solution, causing a concentration gradient that powers a material flow towards the microarray surface. Only labelled strands of the amplicons are captured by probes on the microarray surface and as a result locally the PCR process is not symmetric anymore. Moreover, in course of the process more and more single stranded DNA renatures, leaving relatively less strands and complexes available for hybridization.The main conclusion is that surface fluorescence scales with the bulk concentration of the targets involved. Our analysis shows that the Ct value is only slightly dependent on the initial enzyme load and the degradation of the capture probes. The Ct value, however, does depend strongly on the rate constants of the annealing/hybridization reactions in the bulk and on surface. Local asymmetry appears to be a minor effect.

Effects of cationic and anionic nanoparticles on the stability of the secondary structure of DNA

The effects of anionic and cationic nanoparticles (15 and 100 nm in diameter) on the stability of the secondary structure of DNA were studied in DNA denaturation and renaturation processes. The results showed that DNA denaturation is greatly inhibited in the presence of cationic nanoparticles. In contrast, in the presence of even large amounts of anionic nanoparticles, the DNA melting transition occurs at the original melting temperature. The effects of the size and concentration of nanoparticles on DNA denaturation are discussed.

Electrochemically-Driven Large Amplitude pH Cycling for Acid–Base Driven DNA Denaturation and Renaturation

In this paper, we present an electrochemically driven large amplitude pH alteration method based on a serial electrolytic cell involving a hydrogen permeable bifacial working electrode such as Pd thin foil. The method allows solution pH to be changed periodically up to ±4~5 units without additional alteration of concentration and/or composition of the system. Application to the acid-base driven cyclic denaturation and renaturation of 290 bp DNA fragments is successfully demonstrated with in situ real-time UV spectroscopic characterization. Electrophoretic analysis confirms that the denaturation and renaturation processes are reversible without degradation of the DNA. The serial electrolytic cell based electrochemical pH alteration method presented in this work would promote investigations of a wide variety of potential-dependent processes and techniques.

Improved procedure for the measurement of telomere length in whole cells by PNA probe and flow cytometry

Peptide nucleic acid (PNA) probes hybridize to denatured telomeric sequences in cells permeabilized in hot formamide. In reported protocols, the hybridization was conducted in solutions with high formamide concentrations to avoid the DNA renaturation that can hamper binding of the oligo-PNA probe to specific sequences. We postulated that telomeric DNA, confined in the nuclear microvolume, is not able to properly renature after hot formamide denaturation. Therefore, to improve hybridization conditions between the probe and the target sequences, it might be possible to add probe to sample after the complete removal of formamide. After telomeric DNA denaturation in hot formamide solution and several washes to remove the ionic solvent, cells were hybridized overnight at room temperature with human telomere-specific PNA probe conjugated with Cy5 fluorochrome, Cy5-OO-(CCCTAA)(3) . After stringency washes and staining with ethidium bromide, the cells were analysed by flow cytometry and by using a confocal microscope. Using three continuous cell lines, different in DNA content and telomere length, and resting human peripheral blood T and B lymphocytes, we demonstrated that the oligo-PNA probe hybridized to telomeric sequences after complete removal of formamide and that in the preserved nucleus, telomeric sequence denaturation is irreversible. According to our experience, oligo-PNA binding results is efficient, specific and proportional to telomere length. These, our original findings, can form the technological basis of actual in situ hybridization on preserved whole cells.

Structural modeling of Gene 32 protein and SSB's roles in DNA replication, recombination and repair

Students participating in the Students Modeling a Research Topic (SMART) team partnership program involving Lackey High School, University of Maryland Baltimore County and Milwaukee School of Engineering used 3‐D printing technology to create a physical model of T4 bacteriophage gene 32 protein and researched its role in denaturation and renaturation of DNA. Gene 32 protein is a single‐stranded binding protein (SSB) that has three domains: the core domain with an OB fold provides a structural trough for binding single stranded DNA (ssDNA); the B domain (amino acids 1–21) is responsible for cooperatively interacting with the core domains of other SSBs; and the A domain (C‐terminal 48 amino acids) binds to other proteins of the replication machinery and acts as a gate keeper by mimicking DNA thus controlling DNA binding. Gene 32 protein binds weakly to double stranded DNA (dsDNA). It is theorized that a region on the protein surface rich in lysine residues electrostatically interacts with the phosphates in the dsDNA major groove. dsDNA interaction might provide a means by which the protein could rapidly one‐dimensionally diffuse to transient single stranded regions on the DNA, thus facilitating its roles in DNA replication, recombination and repair. Supported by a grant from HHMI.

Problem-solving test: Dideoxy sequencing

Terms to be familiar with before you start to solve the test: deoxyribonucleoside triphosphates (dNTPs), in vitro DNA synthesis, template, primer, DNA polymerase, denaturation and renaturation of DNA, urea, polyacrylamide gel electrophoresis, and autoradiography.