Gel Electrophoretic Method Research Articles

An easy preparative gel electrophoretic method for targeted depletion of hemoglobin in erythrocyte cytosolic samples

Hemoglobin (Hb) approximately constitutes 98% of the protein composition of a red blood cell (RBC), thus masking the remaining 2% which has still to be discovered completely due to the difficulty in its analysis. Here, we proposed a large-scale native gel electrophoresis that effectively tackles this limitation through a novel sample preparation strategy able to concentrate low-abundance species by removing Hb by means of electrophoretic instruments. Clear native PAGE was performed in a gel electrophoresis tube where the run was intermittently interrupted and different fractions were recovered in liquid phase into a collection chamber placed at the end of the tube. In this way, fractions containing multi-protein complexes with different molecular weights were collected in the native form by a simple elution. Red fraction containing Hb multi-protein complexes can be excluded from subsequent analyses, or rather be analyzed separately, reducing therefore the dynamic range of erythrocyte cytosolic protein concentrations and increasing the number of protein identifications. In particular, 838 protein spots in total were detected when fractions were analyzed by 2-D IEF-SDS-PAGE. This depletion method is inexpensive, simple to perform, reproducible and makes it possible to process large amounts of sample (up to 150 mg), thus making it suitable for in-depth proteome investigations. Furthermore, this strategy has the potential to be applied both to native and denatured proteomes of different biological samples.

Mass Spectrometric Identification of Oxidative Modifications of Tryptophan Residues in Proteins: Chemical Artifact or Post-Translational Modification?

Oxidative modification of tryptophan to kynurenine (KYN) and N-formyl kynurenine (NFK) has been described in mitochondrial proteins associated with redox metabolism, and in human cataract lenses. To a large extent, however, previously reported identifications of these modifications were performed using peptide mass fingerprinting and/or tandem-MS data of proteins separated by gel electrophoresis. To date, it is uncertain whether NFK and KYN may represent sample handling artifacts or exclusively post-translational events. To address the problem of the origin of tryptophan oxidation, we characterized several antibodies by liquid chromatography-tandem mass spectrometry, with and without the use of electrophoretic separation of heavy and light chains. Antibodies are not normally expected to undergo oxidative modifications, however, several tryptophan (Trp) residues on both heavy and light chains were found extensively modified to both doubly oxidized Trp and KYN following SDS-PAGE separation and in-gel digestion. In contrast, those residues were observed as non-modified upon in-solution digestion. These results indicate that Trp oxidation may occur as an artifact in proteins separated by SDS-PAGE, and their presence should be carefully interpreted, especially when gel electrophoretic separation methods are employed.

Angiopoietin-2 in experimental colitis

The pathophysiology of inflammatory bowel disease (IBD) includes leukocyte infiltration, blood and lymphatic remodeling, weight loss and protein enteropathy. The roles of angiopoietin-2 (Ang-2) in initiating gut inflammation, leukocyte infiltration and angiogenesis are not well understood. Disease activity index, histopathological scoring, myeloperoxidase assay, immunohistochemistry and sodium dodecyl sulphate- polyacrylamide gel electrophoretic methods were employed in the present study to address the roles of Ang-2 in experimental colitis. Several important differences were seen in the development of experimental IBD in Ang-2(-/-) mice. Although weight change and disease activity differ only slightly in WT and Ang-2(-/-) + DSS treated mice, leukocyte infiltration, inflammation and blood and lymphatic vessel density is significantly attenuated compared to WT + DSS mice. Gut capillary fragility and water export (stool blood and form) appear significantly earlier in Ang-2(-/-) + DSS mice vs. WT. Colon lengths were also significantly reduced in Ang-2(-/-) and gut histopathology was less severe in Ang-2(-/-) compared to WT + DSS. Lastly, the decrease in serum protein content in WT + DSS was less severe in Ang-2(-/-) + DSS, thus protein losing enteropathy (PLE) a feature of IBD is relieved by Ang-2(-/-). These data demonstrate that in DSS colitis, Ang-2 mediates inflammatory hemangiogenesis, lymphangiogenesis and neutrophil infiltration to reduce some, but not all clinical features of IBD. The implications for Ang-2 manipulation in the development of IBD and other inflammatory diseases and treatments involving Ang-2 are discussed.

Thermodynamics of Forming a Parallel DNA Crossover

The process of genetic recombination involves the formation of branched four-stranded DNA structures known as Holliday junctions. The Holliday junction is known to have an antiparallel orientation of its helices, i.e., the crossover occurs between strands of opposite polarity. Some intermediates in this process are known to involve two crossover sites, and these may involve crossovers between strands of identical polarity. Surprisingly, if a crossover occurs at every possible juxtaposition of backbones between parallel DNA double helices, the molecules form a paranemic structure with two helical domains, known as PX-DNA. Model PX-DNA molecules can be constructed from a variety of DNA molecules with five nucleotide pairs in the minor groove and six, seven or eight nucleotide pairs in the major groove. A topoisomer of the PX motif is the juxtaposed JX1 molecule, wherein one crossover is missing between the two helical domains. The JX1 molecule offers an outstanding baseline molecule with which to compare the PX molecule, so as to measure the thermodynamic cost of forming a crossover in a parallel molecule. We have made these measurements using calorimetric and ultraviolet hypochromicity methods, as well as denaturing gradient gel electrophoretic methods. The results suggest that in relaxed conditions, a system that meets the pairing requirements for PX-DNA would prefer to form the PX motif relative to juxtaposed molecules, particularly for the 6:5 structure.

Electrodeposition of lead zirconate titanate nanotubes

Lead zirconate titanate (PZT) nanotubes have been grown using porous anodic alumina templates. Sol–gel electrophoretic deposition method was utilized to form the nanotubes on pore walls. The templates were prepared using various anodizing voltages to achieve different pore diameters. Phosphoric acid solution was employed as the electrolyte. Stabilized PZT sols were prepared using lead acetate trihydrate and modified precursors of zirconium and titanium with acetic acid. The filled templates were then sintered at 700 °C. Scanning electron microscopy (SEM) shows that tubular PZT arrays have been efficiently grown in the alumina templates. Transmission electron microscopy (TEM) further confirms the tubular form and polycrystalline nature of the tubes. Energy dispersive X-ray (EDX) analyses also confirm the composition of the tubes. X-ray diffraction (XRD) spectra indicate the presence of the perovskite PZT as the main phase.

Purification and characterization of transcribed RNAs using gel filtration chromatography

RNA synthesis using in vitro transcription by phage T7 RNA polymerase allows preparation of milligram quantities of RNA for biochemical, biophysical and structural investigations. Previous purification approaches relied on gel electrophoretic or gravity-flow chromatography methods. We present here a protocol for the in vitro transcription of RNAs and subsequent purification using fast-performance liquid chromatography. This protocol greatly facilitates production of RNA in a single day from transcription to purification.

Comparison of planar SDS-PAGE, CGE-on-a-chip, and MALDI-TOF mass spectrometry for analysis of the enzymatic de-N-glycosylation of antithrombin III and coagulation factor IX with PNGase F

Three different analytical techniques (planar SDS-PAGE, CGE-on-a-chip and MALDI-TOF-MS) applied for determination of the molecular weight of intact and partly and completely de-N-glycosylated human serum glycoproteins (antithrombin III and coagulation factor IX) have been compared. N-Glycans were removed from the protein backbone of both complex glycoproteins using PNGase F, which cleaves all types of asparagine-attached N-glycan provided the oligosaccharide has at least the length of a chitobiose core unit. Two of the applied techniques were based on gel electrophoretic separation in the liquid phase while the third technique was the gas-phase technique mass spectrometry. It was demonstrated that the enzymatic de-N-glycosylation generally worked well (completely or partially) with both glycoproteins (one containing only N-glycans and the second N- and O-glycans). All three methods were suitable for monitoring the de-N-glycosylation progress. While the molecular weights determined with MALDI-TOF-MS were most accurate, both gel electrophoretic methods provided molecular weights that were too high because of the attached glycan structures.

Capillary Electrophoresis Is Essential for Microsatellite Marker Based Detection and Quantification of Adulteration of Basmati Rice ( Oryza sativa)

Microsatellite markers are employed for genotyping of Basmati varieties and assaying purity of market samples. However, employment of diverse electrophoresis techniques across laboratories has resulted in inconsistent allele sizes, creating doubts about the suitability of the assay. This study evaluated agarose gel electrophoresis, slab gel electrophoresis, and capillary electrophoresis techniques for their efficiency in the detection and quantification of adulteration in Basmati samples. Comparative analysis across 8 microsatellite loci in 12 rice varieties demonstrated that the capillary electrophoresis method showed less error (+/-0.73 bp) in the estimation of allele sizes compared to slab gel (+/-1.59 bp) and agarose gel (+/-8.03 bp) electrophoretic methods. Capillary electrophoresis showed greater reproducibility (<0.5 bp deviation) compared to slab gel (1 bp) and agarose (>3 bp) based methods. Capillary electrophoresis was significantly superior in quantification of the adulterant, with a mean error of +/-3.91% in comparison to slab gel (+/-6.09%). Lack of accuracy and consistency of the slab gel and agarose electrophoretic methods warrants the employment of capillary electrophoresis for Basmati rice purity assays.

Synthesis, magnetic property and DNA cleavage behavior of a new dialkoxo‐bridged diiron(III) complex

AbstractA new dialkoxo‐bridged diiron(III) complex, [Fe2(BMA)2(CH3O)2Cl2]·2Cl·4CH3OH (1) [BMA = N,N‐bis(2‐benzimidazolylmethyl)amine], was synthesized and characterized by UV‐visible absorption and infrared spectra and magnetic susceptibilities. The complex crystallizes in the monoclinic system, space group P2(1)/n, a = 12.9659(19) Å, b = 10.0278(16) Å, c = 17.919(2) Å, β = 93.766(8)° , V = 2324.8(6) Å3, Z = 2, F(000) = 1036, Dc = 1.426 g cm−3, µ = 0.908 mm−1. According to X‐ray crystallographic studies, each Fe(III) ion lies in a highly distorted octahedral environment, and two Fe(III) ions are bridged by the methoxyl oxygens. Cryomagnetic analyses indicated a moderate antiferromagnetic interaction between the high‐spin Fe(III) ions, with J = − 27.05 cm−1. Moreover, the binding interaction of DNA with the diiron complex was investigated by spectroscopic and agarose gel electrophoretic methods, showing moderate cleavage activity on pBR322 plasmid DNA at physiological pH and temperature. Copyright © 2007 John Wiley &amp; Sons, Ltd.

Analysis of the synaptic vesicle proteome using three gel-based protein separation techniques

Synaptic vesicles are key organelles in neurotransmission. Their functions are governed by a unique set of integral and peripherally associated proteins. To obtain a complete protein inventory, we immunoisolated synaptic vesicles from rat brain to high purity and performed a gel-based analysis of the synaptic vesicle proteome. Since the high hydrophobicity of integral membrane proteins hampers their resolution by gel electrophoretic techniques, we applied in parallel three different gel electrophoretic methods for protein separation prior to MS. Synaptic vesicle proteins were subjected to either 1-D SDS-PAGE along with nano-LC ESI-MS/MS or to the 2-D gel electrophoretic techniques benzyldimethyl-n-hexadecylammonium chloride (BAC)/SDS-PAGE, and double SDS (dSDS)-PAGE in combination with MALDI-TOF-MS. We demonstrate that the combination of all three methods provides a comprehensive survey of the proteinaceous inventory of the synaptic vesicle membrane compartment. The identified synaptic vesicle proteins include transporters, soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs), synapsins, rab and rab-interacting proteins, additional guanine nucleotide triphosphate (GTP) binding proteins, cytoskeletal proteins, and proteins modulating synaptic vesicle exo- and endocytosis. In addition, we identified novel proteins of unknown function. Our results demonstrate that the parallel application of three different gel-based approaches in combination with mass spectrometry permits a comprehensive analysis of the synaptic vesicle proteome that is considerably more complex than previously anticipated.

Template-directed formation of functional complex metal-oxide nanostructures by combination of sol–gel processing and spin coating

We report the template-based formation of functional complex metal-oxide nanostructures by a combination of sol–gel processing and spin coating. This method employs the spin-coating of a sol–gel solution into an anodic aluminum oxide membrane (SSAM). Various metal-oxide nanowires and nanotubes with a high aspect-ratio were prepared. The aspect-ratios of the PbO 2 nanowires and Pb(Zr 0.52Ti 0.48)O 3 nanowires were about 300 and 400, respectively, and their diameters were about 50 nm. The fabricated PbTiO 3 nanotubes have a relatively constant wall thickness of about 20 nm with an outer diameter of about 60 nm. The deposition time for all of the fabricated metal-oxide nanowires and nanotubes is less than 120 s, which is far shorter than those required in both the sol–gel dipping and sol–gel electrophoretic methods. These results indicate that the SSAM method can be a versatile pathway to prepare functional complex metal-oxide nanowires and nanotubes with a high aspect-ratio. The possible formation process for the one-dimensional nanostructures by SSAM is discussed.

Single-based resolution for oligodeoxynucleotides and their phosphorothioate modifications by replaceable capillary gel electrophoresis

A replaceable capillary gel electrophoretic (replaceable CGE) method was developed for the separation of two sets of model compounds of single-stranded oligodeoxynucleotide mixtures (18-20 mers), phosphodiester oligodeoxynucleotides (PO-ODNs) and their phosphorothioate modifications (PS-ODNs), with equal sequences differing in a single base. Polyethylene glycol (PEG) 35000 was chosen as the sieving matrix. It was confirmed that PEG polymer solution less influenced resolutions of the PS-ODNs compared with those of the PO-ODNs, while acetonitrile used as an additive in the system improved the separation significantly. It was also noticed that the effect of temperature on separation was much larger than that of denaturant urea.

Changes of nuclear matrix protein and its relationship with c-erbB-2 in human colon adenocarcinoma

Objective: Nuclear matrix protein is tissue, cell-type specific, and tumor-relative. It plays an important role in the regulation of intranuclear processes. Some researches also showed that a c-erbB-2 promoter-specific DNA-binding nuclear matrix protein is present only in malignant human breast tissues and induces mitogenesis and cell surface expression of the c-erbB-2 protein in resting NIH/3T3 cells. But it is not clear that how it in colon adenocarcinomas. Methods: Two-dimensional gel electrophoretic method was used for NMP identification and immunohistochemistry was used for c-erbB-2 detection in 12 cases of colon adenocarcinomas and matched adjacent normal colon tissues. Results: 5 different nuclear matrix proteins (named C1–C5) were identified in 12 colon adenocarcinoma specimens, but not in the matched adjacent normal colon tissues; 3 nuclear matrix proteins (named N1–N3) were identified in all 12 matched adjacent normal colon tissues, but not in colon adenocarcinoma specimens. A nuclear matrix protein (named N4) was detected in all of 9 moderated-well differentiated adenocarcinomas and all 12 matched adjacent normal colon tissues, but not in 3 poor-differentiated adenocarcinomas. All of the 10 colon adenocarcinomas which had the nuclear matrix protein C4 were c-erbB-2 expression positive. Conclusion: The data suggest that there are specific nuclear matrix proteins in colon adenocarcinomas and its subtypes, which maybe valuable to serve as markers of colon adenocarcinomas in future. Nuclear matrix protein C4 probably is a c-erbB-2 promotor-specific nuclear matrix protein in colon adenocarcinomas, and may induce the expression of c-erbB-2.

Topological measurement of an A-tract bend angle: effect of magnesium.

Sequences of four to six adenine residues, termed A-tracts, have been shown to produce curvature in the DNA double helix. A-tracts have been used extensively as reference standards to quantify bending induced by other sequences as well as by DNA binding proteins when they bind to their sites. However, the ability of an A-tract to serve as such a standard is hampered by the wide variation of values reported for the amount of bend conferred by an A-tract. One experimental condition that differs in these studies is the presence of divalent cation. To evaluate this effect, a new application of a topological method, termed rotational variant analysis, is used here to measure for the first time the effect of the presence of magnesium ion on the bend angle conferred by an A-tract. This method, which has the unique ability to measure a bend angle in the presence or absence of magnesium ion, demonstrates that magnesium ion markedly increases the bend angle. For example, when measured in a commonly used gel electrophoretic buffer, the bend angle conferred by a tract of six adenine residues increases from about 7 degrees in the absence of magnesium ion to 19 degrees in the presence of 3.9 mM magnesium ion. This quantitative demonstration of substantial magnesium ion dependence has several important implications. First, it explains discrepancies among bend values reported in various previous studies, particularly those employing gel electrophoretic versus other solution methods. In addition, these findings necessitate substantial revisions of the conclusions in a large number of studies that have used A-tract DNA as the bend angle reference standard in comparison measurements. Finally, any such future studies employing this comparison methodology will need to use the same sequence analyzed in the original measurements as well as replicate the original measurement conditions (e.g. ionic composition and temperature).

Multiple competitive PCR-DGGE as a tool for quantifying and profiling defined mixed cultures of lactic acid bacteria during production of probiotics from complex polysaccharides.

To apply a denaturing gradient gel electrophoretic (DGGE) method to quantify and profile individual strains during a mixed culture fermentation. DNA was extracted during the culture of lactic acid bacteria (LAB) and amplified in a multiple competitive PCR (cPCR) using general primers targeting 16S rDNA and DNA from Streptococcus salivarius as competitive DNA. Subsequently the 200-kb amplified fragments were separated by DGGE. The method was validated in pure cultures and used to profile a mixture of three LAB grown on glucose, soluble starch and glycogen from mussel processing waste. The inclusion of a starch- and glycogen-degrading strain (Lactobacillus plantarum) and a weakly amylotic nisin-resistant strain (Lact. paracasei) allowed proliferation of the nisin producing Lactococcus lactis which in itself is unable to grow on complex carbohydrates. cPCR-DGGE permitted the monitoring of a different strain succession on the different carbohydrates, related to amylolytic activity and substrate consumption, acid production and nisin production. cPCR-DGGE is a useful tool for profiling defined mixed cultures of bacteria and hence allows their interaction to be studied. Provided validation of the method for each specific case, it may be appropriate to monitor and control the reproducibility of any defined mixed culture of bacteria, with the advantage that an increase in the strain numbers to be monitored does not yield an increase in the labour of the procedure.

Detection of alcohol dehydrogenase isoenzymes in liver and serum by electrophoresis in liver disease

AIM: To establish an agarose gel electrophoretic method for detecting alcohol dehydrogenase isoenzymes in human serum or liver tissue. METHODS: The samples of human liver tissue or serum were electrophoresed with 74 mmol/L diethylbarbital buffer (pH 8.6) of 10 g/L agarose gel. Electrophoresis can separate the Alcohol Dehydrogenase (ADH) isoenzymes to three bands clearly at 20 mA for 20 min in serum. RESULTS: The total ADH activity in serum of sixty-seven patients with liver diseases was ranged from 0.013 Kat/L to 0.021 Kat/L. ADH isoenzyme was ranged from 0.01 to 0.30 of the total activities . ADH isoenzyme was from 0.12 to 0.31 . ADH was from 0.39 to 0.80. Total ADH activity in liver tissues of ten healthy subjects was ranged from 0.136 Kat/L to 0.196 Kat/L. ADH isoenzyme was from 0.07 to 0.25 of the total activities. ADH isoenzyme was ranged from 0.19 to 0.27. ADH was from 0.56 to 0.73. CONCLUSION: ADH activity is high in normal human liver. Three ADH isoenzymes can be separated with agarose gel electrophoresis . But serum ADH activity is low. High activities were obtained in serum from persons suffering from serious liver diseases. Mi QM, Cao LN, Gao CF. Detection of alcohol dehydrogenase isoenzymes in liver and serum by electrophoresis in liver disease. Shijie Huaren Xiaohua Zazhi 2003;11(8):1175-1177

Allelic variation of the HMW glutenin subunits in Aegilops tauschii accessions detected by sodium dodecyl sulphate (SDS-PAGE), acid polyacrylamide gel (A-PAGE) and capillary electrophoresis

Variability of high molecular weight glutenin subunits (HMW-GS) was studied in198 accessions of Ae. Tauschii (2n=2x=14, DD) by sodium dodecyl sulphate(SDS-PAGE) and acid polyacrylamide gel electrophoresis (A-PAGE) and capillary electrophoresis (CE). A high allelic variation of HMW-GS, including some novel x- and y-type subunits and variable subunit combinations were observed. One accession(TD159) showed a x-type null form. The results by A-PAGE analysis revealed that the subunits Dx5 t and Dy10 t encoded by Glu-D t 1 locus in Ae. tauschii were different in relative mobilities in comparison with the subunits Dx5 and Dy10 found in bread wheats, whereas they had the same mobilities, respectively, when separated by SDS-PAGE. The higher resolution of Ae. tauschii HMW-GS separated by CE method showed two clear peaks in accordance with x- and y-type subunits, respectively,except the accession TD151 which possessed only subunit Dy12.1*t. The electro elution time of the x-type and y-type subunits were about 13–14 and 7–8minutes, respectively. Characterization of wheat HMW-GS was facilitated by using CE which provides high resolution and increases the speed of analysis in conjunction with the traditional gel electrophoretic methods. A total of 42HMW-GS alleles were identified, among which were several alleles not presently detected in bread wheats. Hence Ae. tauschii is potentially a valuable genetic resource for quality improvement of bread wheat.

THE USE OF GEL ELECTROPHORESIS AND IMMUNOLOGICAL TECHNIQUES IN THE ANALYSIS OF GUM ARABIC

Gum arabic, also known as acacia gum, is an abundant natural product that has been used throughout history. It has been extensively used in the manufacture of eastern and western inks, watercolors and gouache as a binder, and in historic photographic processes such as gum bichromate prints. Much research has been done on the analysis of acacia gums yet the main obstacle to their characterization has been the absence of an efficient separation technique. The most widely used technique has been size exclusion/gel filtration chromatography (SEC/GFC); however, even GFC has not provided adequate separation of the gum components. In addition to a brief overview of acacia gums, this presentation reviews the development of a new gel electrophoretic method for the separation and detection of the polysaccharides, glycoproteins and proteins that constitute acacia gums, and the use of specific antibodies to identify acacia gums at the species level.

Genetic analysis of sea-ice bacterial communities of the Western Baltic Sea using an improved double gradient method

The bacterial diversity of sea ice from Kiel Bight obtained during the rare event of solid ice cover in spring 1996 was analysed by molecular genetic approaches using an improved double gradient denaturing gradient gel electrophoretic method (DG-DGGE) to separate 16S rDNA fragments of approximately 500 bp. The excellent separation of individual bands within these gradient gels allowed us to obtain sequence information and to allocate the phylogenetic position of representative bacteria from the sea ice. The band pattern of the gradient gels revealed a vertical stratification of the bacterial species distribution within the ice and the presence of characteristic bacteria for each layer. According to their 16S rDNA sequences, major bands of the gradient gels represented bacteria closely related to fermenting species of the genera Propionibacterium and Bacteroides and to anoxygenic phototrophic purple sulfur bacteria (Chromatiaceae). Their abundance in horizons of the inner ice core may indicate the existence of oxygen-deficient and anoxic zones or niches and possible primary production by anoxygenic photosynthesis within the investigated Baltic Sea sea ice. This is the first phylogenetic evidence of the presence, and most probably the development, of phototrophic purple sulfur bacteria in sea ice.

DNA-bending finger: artificial design of 6-zinc finger peptides with polyglycine linker and induction of DNA bending.

DNA structural changes such as bending play an important role in various biological reactions. Not only protein binding to its specific DNA sequence but also DNA bending induced by the protein is indispensable for unique gene expression. Therefore, an artificial protein that induces a DNA conformational change is interesting as a transcriptional regulator of a specific gene. We created 6-zinc finger proteins, Sp1ZF6(Gly)n (n = 4, 7, 10), by connecting two DNA binding domains of transcription factor Sp1 with flexible polyglycine peptide linkers, and their effects on DNA structure were compared with that of native 3-zinc finger Sp1(530-623). Gel electrophoretic methods revealed that Sp1ZF6(Gly)7 and Sp1ZF6(Gly)10 bind to two distal GC boxes and result in DNA bending. Evidently, the hydroxyl radical footprinting analysis demonstrated that hypersensitive cleavage was observed at the 5'-TA step in the intervening region bound by Sp1ZF6(Gly)7 or Sp1ZF6(Gly)10. The phasing assays strongly suggested that the induced DNA bending was directed toward the major groove and that Sp1ZF6(Gly)7 caused the most drastic directional change in DNA bending. Of special interest are the facts that the newly designed 6-finger peptides Sp1ZF6(Gly)7 and Sp1ZF6(Gly)10 can induce DNA bending at the intervening region of the two distal binding sites and that the linker length between two 3-zinc finger motifs has a crucial effect on the entire DNA-bending direction. Such DNA-bending fingers may be feasible for use as a gene expression regulator based on the structural change in DNA in the future.