Characterisation of optically driven microstructures for manipulating single DNA molecules under a fluorescence microscope

Optical tweezers are powerful tools for manipulating single DNA molecules using fluorescence microscopy, particularly in nanotechnology‐based DNA analysis. We previously proposed a manipulation technique using microstructures driven by optical tweezers that allows the handling of single giant DNA molecules of millimetre length that cannot be manipulated by conventional techniques. To further develop this technique, the authors characterised the microstructures quantitatively from the view point of fabrication and efficiency of DNA manipulation under a fluorescence microscope. The success rate and precision of the fabrications were evaluated. The results indicate that the microstructures are obtained in an aqueous solution with a precision ∼50 nm at concentrations in the order of 10⁶ particles/ml. The visibility of these microstructures under a fluorescence microscope was also characterised, along with the elucidation of the fabrication parameters needed to fine tune visibility. Manipulating yeast chromosomal DNA molecules with the microstructures illustrated the relationship between the efficiency of manipulation and the geometrical shape of the microstructure. This report provides the guidelines for designing microstructures used in single DNA molecule analysis based on on‐site DNA manipulation, and is expected to broaden the applications of this technique in the future.Inspec keywords: DNA, molecular biophysics, fluorescence, optical microscopy, radiation pressure, biological techniquesOther keywords: optically driven microstructures, single DNA molecule analysis, fluorescence microscopy, optical tweezers, nanotechnology‐based DNA analysis, manipulation technique, aqueous solution, fine tune visibility, yeast chromosomal DNA molecules, geometrical shape, on‐site DNA manipulation     

Increase in weld penetration depth by two Nd: YAG laser beams combined

Summary

As part of current R&;D work focused on developing the intelligence of arc welding robots, this paper deals with the problem of weldpool sensing and control. To obtain high‐quality welding, it is important to control the weldpool depth in robot welding regardless of any external disturbance, such as irregular groove gap. The method of controlling the weldpool depth without a mathematical model is discussed.

Since it is difficult to measure the weldpool depth directly, it is estimated from the weldpool surface shape, groove gap, and welding current. A neural network is used to estimate the weldpool depth without a mathematical model. The weldpool depth is controlled from the output of the neural network using the fuzzy controller. Neural network and fuzzy controller application is validated in welding experiments.     

Conditional transformation of immunoglobulin mutation pattern from gene conversion into point mutation by controlling XRCC3 expression in the DT40 B cell line

A hypermutating B cell line DT40 is useful for screening antibodies and improving affinity of the selected antibodies in vitro. To perform affinity maturation efficiently, we generated an engineered DT40 line whose immunoglobulin mutation pattern can be transformed from gene conversion into point mutation by conditional suppression of XRCC3 expression.     

Interfacial microstructure and strength of steel/aluminum alloy joints welded by resistance spot welding with cover plate

We joined aluminum alloy A5052 to cold-rolled steel SPCC (Steel Plate Cold Commercial) and austenitic stainless steel SUS304 using resistance spot welding with a cover plate. The interfacial microstructure was observed using transmission electron microscopy. A thick two-layered reaction layer contains Fe₂Al₅ and FeAl₃ and a thin serration reaction layer contains Fe₂Al₅ and FeAl₃ were observed at the A5052/SPCC and A5052/SUS304 interface, respectively. Mechanical property analysis suggested that the reaction layer has no effect on the tensile shear strength of the A5052/SUS304 joint and that the tensile shear strength of the A5052/SPCC joint is influenced by the reaction layer formed at its interface.     

Daily variation in an intensity of kaolin-induced writhing reaction in mice

The daily variation in an intensity of kaolin-induced writhing reaction was examined in mice kept under conditions of light; 07:00 - 19:00 and dark; 19:00 - 07:00. The number of writhes was counted for 30 minutes after a single intraperitoneal injection of kaolin at 00:00, 02:00, 04:00, 06:00, 08:00, 10:00, 12:00, 14:00, 16:00, 18:00, 20:00 and 22:00. The number of writhes showed a daily variation with a peak at 18:00 and a trough at 06:00. The intensity of writhing reaction was significantly reduced by pretreatment with the bradykinin B1 (Des-Arg9-[Leu8]-BK) and B2 (icatibant) receptor antagonists. Significant daily variation in this parameter was still observed in the group with the B1 antagonist, but disappeared in the B2 antagonist-treated group. These results suggest that the kaolin-induced writhing reaction shows the daily variation with a peak at the end of the resting period and a trough at the end of the active period. The B2 receptor mediated stimuli appears to be involved in this phenomenon.     

Photoresponse and donor concentration of plasma-sprayed TiO<Subscript>2</Subscript> and TiO<Subscript>2</Subscript>-ZnO electrodes

The photoelectrochemical characteristics of plasma-sprayed porous TiO₂, TiO₂-5%ZnO, and TiO₂-10%ZnO electrodes in 0.1 N NaOH solution were studied through a three-electrode cell system. The microstructure, morphology, and composition of the electrodes were analyzed using an electron probe surface roughness analyzer (ERA-8800FE), scanning electron microscopy, and x-ray diffraction. The results indicate that the sprayed electrodes have a porous microstructure, which is affected by the plasma spray parameters and composition of the powders. The TiO₂-ZnO electrodes consist of anatase TiO₂, rutile TiO₂, and Zn₂Ti₃O₈ phase. The photoresponse characteristics of the plasma-sprayed electrodes are comparable to those of single-crystal TiO₂, but the breakdown voltage is close to 0.5 V (versus that of a saturated calomel electrode). The short-circuit photocurrent density (J _SC) increases with a decrease of donor concentration, which was calculated according to the Gartner-Butler model. For the lowest donor concentration of a TiO₂-5%ZnO electrode sprayed under an arc current of 600 A, the short-circuit J _SC is approximately 0.4 mA/cm² higher than that of the TiO₂ electrodes under 30 mW/cm² xenon light irradiation. The J _SC increases linearly with light intensity. The original version of this paper was published as part of the DVS Proceedings: “Thermal Spray Solutions: Advances in Technology and Application,” International Thermal Spray Conference, Osaka, Japan, 10–12 May 2004, CD-Rom, DVS-Verlag GmbH, Düsseldorf, Germany.     

Towards a local learning (innovation) model of solar photovoltaic deployment

It is by now familiar that in the deployment of solar photovoltaic (PV) systems, the cost dynamics of major system component like solar cell/module is subjected to experience curve effects driven by production learning and research and development at the supplier side. What is less clear, however, is the economics of system integration or system deployment that takes place locally close to the user, involving other market players, in the downstream solar PV value chain. Experts have agreed that suppliers of solar PV system must customize their flexible characteristics to address local unique users’ and applications requirements and compete on price/performance basis. A lack of understanding of the drivers of the economics of system customization therefore is a deficiency in our understanding of the overall economics of this renewable energy technology option.     

Synthesis of ordered mesoporous carbon thin films at various temperatures in vapor infiltration method

Mesoporous carbon thin films with ordered structures were prepared by using resorcinol-surfactant self-assembly. A mixture of resorcinol, surfactant, and ethanol coated on silicon substrates was exposed to formaldehyde vapor as a cross-linking agent to form structured resorcinol/formaldehyde resin films. The films were then carbonized at 800 °C in an inert atmosphere to remove the surfactant and to obtain structured carbon materials. With this vapor infiltration method, thin films with several structures were obtained from the same precursor solution by employing different vapor infiltration temperatures. The results were interpreted from the transformation of the self-assembly during the vapor infiltration process.     

Relationship between Expression of Onco-Related miRNAs and the Endoscopic Appearance of Colorectal Tumors

Accumulating data indicates that certain microRNAs (miRNAs or miRs) are differently expressed in samples of tumors and paired non-tumorous samples taken from the same patients with colorectal tumors. We examined the expression of onco-related miRNAs in 131 sporadic exophytic adenomas or early cancers and in 52 sporadic flat elevated adenomas or early cancers to clarify the relationship between the expression of the miRNAs and the endoscopic morphological appearance of the colorectal tumors. The expression levels of miR-143, -145, and -34a were significantly reduced in most of the exophytic tumors compared with those in the flat elevated ones. In type 2 cancers, the miRNA expression profile was very similar to that of the exophytic tumors. The expression levels of miR-7 and -21 were significantly up-regulated in some flat elevated adenomas compared with those in exophytic adenomas. In contrast, in most of the miR-143 and -145 down-regulated cases of the adenoma-carcinoma sequence and in some of the de novo types of carcinoma, the up-regulation of oncogenic miR-7 and/or -21 contributed to the triggering mechanism leading to the carcinogenetic process. These findings indicated that the expression of onco-related miRNA was associated with the morphological appearance of colorectal tumors.     

Development of a strain for efficient degradation of polychlorinated biphenyls by patchwork assembly of degradation pathways

Rhodococcus jostii RHA1 accumulates chlorobenzoates (CBA) during the degradation of polychlorinated biphenyls (PCBs). CBA degradation is considered one of the rate-limiting steps in the complete degradation of PCBs. To reduce the accumulation of CBAs, the upper pathway enzyme genes for PCB degradation of RHA1 were introduced into a CBA-degrading bacterium, Burkholderia sp. NK8. The resulting recombinant strain exhibited no biphenyl 2,3-dioxygenase (BphA) activity encoded by bphAaAbAcAd genes, which encode the large and small subunits of the terminal oxygenase component and the ferredoxin and reductase subunits responsible for electron transfer from NADH to the large subunit. The remaining enzyme genes involved in the transformation of biphenyl to benzoate, bphB2C1D1, which encode dehydrogenase, ring-cleavage dioxygenase and hydrolase, conferred activities to NK8. To obtain the BphA activity of RHA1 in NK8, sets of BphA genes were constructed by combining the bphAaAbAcAd genes of RHA1 and bphA3A4 of Pseudomonas pseudoalcaligenes KF707, encoding the ferredoxin and reductase subunits. Hybrid derivatives of BphA containing the KF707 bphA3 conferred BphA activity to NK8, and a derivative containing the RHA1 bphAaAb and KF707 bphA3A4 genes exhibited the highest BphA activity. A plasmid containing the RHA1 bphAaAb and KF707 bphA3A4 genes plus the RHA1 bphB2C1D1 genes was constructed and introduced into NK8. The resulting recombinant strain efficiently degraded 2-, 3- and 4-chlorobiphenyls with an apparent reduction in CBA accumulation in comparison to the recombinant mutant strain, which had an insertion in the cbeA gene to inactivate CBA dioxygenase.