A class I odorant receptor enhancer shares a functional motif with class II enhancers.

In the mouse, 129 functional class I odorant receptor (OR) genes reside in a ~ 3 megabase huge gene cluster on chromosome 7. The J element, a long-range cis-regulatory element governs the singular expression of class I OR genes by exerting its effect over the whole cluster. To elucidate the molecular mechanisms underlying class I-specific enhancer activity of the J element, we analyzed the J element sequence to determine the functional region and essential motif. The 430-bp core J element, that is highly conserved in mammalian species from the platypus to humans, contains a class I-specific conserved motif of AAACTTTTC, multiple homeodomain sites, and a neighboring O/E-like site, as in class II OR-enhancers. A series of transgenic reporter assays demonstrated that the class I-specific motif is not essential, but the 330-bp core J-H/O containing the homeodomain and O/E-like sites is necessary and sufficient for class I-specific enhancer activity. Further motif analysis revealed that one of homeodomain sequence is the Greek Islands composite motif of the adjacent homeodomain and O/E-like sequences, and mutations in the composite motif abolished or severely reduced class I-enhancer activity. Our results demonstrate that class I and class II enhancers share a functional motif for their enhancer activity.

Visualization of internal structure and internal stress in visibly opaque objects using full-field phase-shifting terahertz digital holography.

We construct a full-field phase-shifting terahertz digital holography (PS-THz-DH) system by use of a THz quantum cascade laser and an uncooled, 2D micro-bolometer array. The PS-THz-DH enables us to separate the necessary diffraction-order image from unnecessary diffraction-order images without the need for spatial Fourier filtering, leading to suppress the decrease of spatial resolution. 3D shape of a visibly opaque object is visualized with a sub-millimeter lateral resolution and a sub-µm axial resolution. Also, the digital focusing of amplitude image enables the visualization of internal structure with the millimeter-order axial selectivity. Furthermore, the internal stress distribution of an externally compressed object is visualized from the phase image. The demonstrated results imply a possibility for non-destructive inspection of visibly opaque non-metal materials.

Erratum: Author Correction: Bcl11b controls odorant receptor class choice in mice.

[This corrects the article DOI: 10.1038/s42003-019-0536-x.].

Refractive index sensing with temperature compensation by a multimode-interference fiber-based optical frequency comb sensing cavity.

We proposed a refractive index (RI) sensing method with temperature compensation by using an optical frequency comb (OFC) sensing cavity including a multimode-interference (MMI) fiber, namely, the MMI-OFC sensing cavity. The MMI-OFC sensing cavity enables simultaneous measurement of material-dependent RI and sample temperature by decoding from the comb spacing frequency shift and the wavelength shift of the OFC. We realized the simultaneous and continuous measurement of RI-related concentration of a liquid sample and its temperature with precisions of 1.6 × 10-4 RIU and 0.08 °C. The proposed method would be a useful means for the various applications based on RI sensing.

Bcl11b controls odorant receptor class choice in mice.

Each olfactory sensory neuron (OSN) expresses a single odorant receptor (OR) gene from the class I or class II repertoire in mice. The mechanisms that regulate OR class choice in OSNs remain unknown. Here, we show that the transcription factor Bcl11b determines the OR class to be expressed in OSNs. Both loss- and gain-of-function analyses demonstrate that class I is a default fate of OSNs and that Bcl11b dictates a class II OR choice by suppressing the effect of the J-element, a class I-OR enhancer. We further demonstrate that OSN-specific genetic manipulations of Bcl11b bias the OR class choice, generating mice with "class I-dominant" and "class II-dominant" noses, which display contrasting innate olfactory behaviors to two distinct aversive odorants. Overall, these findings reveal a unique transcriptional mechanism mediating a binary switch for OR class choice that is crucial to both the anatomical and functional organization of the olfactory system.

Multicascade-linked synthetic wavelength digital holography using an optical-comb-referenced frequency synthesizer.

Digital holography (DH) is a promising method for non-contact surface topography because the reconstructed phase image can visualize the nanometer unevenness in a sample. However, the axial range of this method is limited to the range of the optical wavelength due to the phase wrapping ambiguity. Although the use of two different wavelengths of light and the resulting synthetic wavelength, i.e., synthetic wavelength DH, can expand the axial range up to several hundreds of millimeters, its axial precision does not reach sub-micrometer. In this article, we constructed a tunable external cavity laser diode phase-locked to an optical frequency comb, namely, an optical-comb-referenced frequency synthesizer, enabling us to generate multiple synthetic wavelengths within the range of 32 µm to 1.20 m. A multiple cascade link of the phase images among an optical wavelength ( = 1.520 µm) and 5 different synthetic wavelengths ( = 32.39 µm, 99.98 µm, 400.0 µm, 1003 µm, and 4021 µm) enables the shape measurement of a reflective millimeter-sized stepped surface with the axial resolution of 34 nm. The axial dynamic range, defined as the ratio of the axial range ( = 2.0 mm) to the axial resolution ( = 34 nm), achieves 5.9 × 105, which is larger than that of previous synthetic wavelength DH. Such a wide axial dynamic range capability will further expand the application field of DH for large objects with meter dimensions.

A Single Pheromone Receptor Gene Conserved across 400 My of Vertebrate Evolution.

Pheromones are crucial for eliciting social and sexual behaviors in diverse animal species. The vomeronasal receptor type-1 (V1R) genes, encoding members of a pheromone receptor family, are highly variable in number and repertoire among mammals due to extensive gene gain and loss. Here, we report a novel pheromone receptor gene belonging to the V1R family, named ancient V1R (ancV1R), which is shared among most Osteichthyes (bony vertebrates) from the basal lineage of ray-finned fishes to mammals. Phylogenetic and syntenic analyses of ancV1R using 115 vertebrate genomes revealed that it represents an orthologous gene conserved for >400 My of vertebrate evolution. Interestingly, the loss of ancV1R in some tetrapods is coincident with the degeneration of the vomeronasal organ in higher primates, cetaceans, and some reptiles including birds and crocodilians. In addition, ancV1R is expressed in most mature vomeronasal sensory neurons in contrast with canonical V1Rs, which are sparsely expressed in a manner that is consistent with the "one neuron-one receptor" rule. Our results imply that a previously undescribed V1R gene inherited from an ancient Silurian ancestor may have played an important functional role in the evolution of vertebrate vomeronasal organ.

Refractive-index-sensing optical comb based on photonic radio-frequency conversion with intracavity multi-mode interference fiber sensor.

Optical frequency combs (OFCs) have attracted attention as optical frequency rulers due to their tooth-like discrete spectra together with their inherent mode-locking nature and phase-locking control to a frequency standard. Based on this concept, their applications until now have been demonstrated in the fields of optical frequency metrology. However, if the utility of OFCs can be further expanded beyond their application by exploiting new aspects of OFCs, this will lead to new developments in optical metrology and instrumentation. Here, we report a fiber sensing application of OFCs based on a coherent link between the optical and radio frequencies, enabling high-precision refractive index measurement based on frequency measurement in radio-frequency (RF) region. Our technique encodes a refractive index change of a liquid sample into a repetition frequency of OFC by a combination of an intracavity multi-mode-interference fiber sensor and wavelength dispersion of a cavity fiber. Then, the change in refractive index is read out by measuring the repetition frequency in RF region based on a frequency standard. Use of an OFC as a photonic RF converter will lead to the development of new applications in high-precision fiber sensing with the help of functional fiber sensors and precise RF measurement.

Scan-less hyperspectral dual-comb single-pixel-imaging in both amplitude and phase.

We have developed a hyperspectral imaging scheme that involves a combination of dual-comb spectroscopy and Hadamard-transform-based single-pixel imaging. The scheme enables us to obtain 12,000 hyperspectral images of amplitude and phase at a spatial resolution of 46 µm without mechanical scanning. The spectral resolution given by the data point interval in the frequency domain is 20 MHz and the comb mode interval is 100 MHz over a spectral range of 1.2 THz centered at 191.5 THz. As an initial demonstration of our scheme, we obtained spectroscopic images of a standard test chart through an etalon plate. The thickness of an absorptive chromium-coated layer on a float-glass substrate was determined to be 70 nm from the hyperspectral phase images in the near-infrared wavelength region.

A long-range cis-regulatory element for class I odorant receptor genes.

Individual olfactory sensory neurons express a single odorant receptor gene from either class I genes residing in a single cluster on a single chromosome or class II genes spread over multiple clusters on multiple chromosomes. Here, we identify an enhancer element for mouse class I genes, the J element, that is conserved through mammalian species from the platypus to humans. The J element regulates most class I genes expression by exerting an effect over ~ 3 megabases within the whole cluster. Deletion of the trans J element increases the expression frequencies of class I genes from the intact J allele, indicating that the allelic exclusion of class I genes depends on the activity of the J element. Our data reveal a long-range cis-regulatory element that governs the singular class I gene expression and has been phylogenetically preserved to retain a single cluster organization of class I genes in mammals."Each olfactory sensory neuron expresses a single odorant receptor gene from either class I or class II genes. Here, the authors identify an enhancer for mouse class I genes, that is highly conserved, and regulates most class I genes expression by acting over ~ 3 megabases within the whole cluster."

Dual-comb spectroscopic ellipsometry.

Spectroscopic ellipsometry is a means of investigating optical and dielectric material responses. Conventional spectroscopic ellipsometry is subject to trade-offs between spectral accuracy, resolution, and measurement time. Polarization modulation has afforded poor performance because of its sensitivity to mechanical vibrational noise, thermal instability, and polarization-wavelength dependency. We combine spectroscopic ellipsometry with dual-comb spectroscopy, namely, dual-comb spectroscopic ellipsometry. Dual-comb spectroscopic ellipsometry (DCSE). DCSE directly and simultaneously obtains the ellipsometric parameters of the amplitude ratio and phase difference between s-polarized and p-polarized light signals with ultra-high spectral resolution and no polarization modulation, beyond the conventional limit. Ellipsometric evaluation without polarization modulation also enhances the stability and robustness of the system. In this study, we construct a polarization-modulation-free DCSE system with a spectral resolution of up to 1.2 × 10-5 nm throughout the spectral range of 1514-1595 nm and achieved an accuracy of 38.4 nm and a precision of 3.3 nm in the measurement of thin-film samples.Spectroscopic ellipsometry is an established technique to characterize the optical properties of a material. Here, Minamikawa et al. combine the method with dual-comb spectroscopy, which allows them to obtain ellipsometric parameters including the phase difference between s-polarized and p-polarized light.

Optical-rotatory-dispersion measurement approach using the nonlinear behavior of the geometric phase.

We propose a method for high-sensitivity optical rotatory dispersion (ORD) measurement of optically active samples that takes advantage of the nonlinear behavior of the geometric phase (GP). To measure the GP as a function of wavelength, we use a multichannel Fourier transform spectrometer (MC-FTS) that is based on Savart plate birefringent-polarization interference, into which we newly insert a zeroth-order quarter-wave plate (QWP). The modified MC-FTS allows us to measure the wavelength dependence of the GP and thus that of the optical rotation angle due to the sample. In this paper, we describe the proposed approach and demonstrate proof-of-principle experiments.

Subpixel-shift cyclic-Hadamard microscopic imaging using a pseudo-inverse-matrix procedure.

We propose two subpixel-shift (SPS) imaging methods: a normal SPS (NSPS) and an interleaved SPS (ISPS) method. Both methods use a series of cyclic-Hadamard-mask illuminations and a single channel detector that enhances lateral spatial resolution in proportion to the number of shifts. A sample object (or the illumination mask) is moved in two dimensions in micro-step intervals within the range of an illumination mask unit pixel (or within a range that exceeds it). To reconstruct the original image, we use a pseudo-inverse-matrix procedure that ensures measurement precision in a least-squares sense. Numerical simulation and experimental results obtained using a transmission microscope are shown. Further preliminary results indicate that the SPS method has the potential to enable measurements exceeding the diffraction limit of optics.

Dynamic terahertz spectroscopy of gas molecules mixed with unwanted aerosol under atmospheric pressure using fibre-based asynchronous-optical-sampling terahertz time-domain spectroscopy.

Terahertz (THz) spectroscopy is a promising method for analysing polar gas molecules mixed with unwanted aerosols due to its ability to obtain spectral fingerprints of rotational transition and immunity to aerosol scattering. In this article, dynamic THz spectroscopy of acetonitrile (CH3CN) gas was performed in the presence of smoke under the atmospheric pressure using a fibre-based, asynchronous-optical-sampling THz time-domain spectrometer. To match THz spectral signatures of gas molecules at atmospheric pressure, the spectral resolution was optimized to 1 GHz with a measurement rate of 1 Hz. The spectral overlapping of closely packed absorption lines significantly boosted the detection limit to 200 ppm when considering all the spectral contributions of the numerous absorption lines from 0.2 THz to 1 THz. Temporal changes of the CH3CN gas concentration were monitored under the smoky condition at the atmospheric pressure during volatilization of CH3CN droplets and the following diffusion of the volatilized CH3CN gas without the influence of scattering or absorption by the smoke. This system will be a powerful tool for real-time monitoring of target gases in practical applications of gas analysis in the atmospheric pressure, such as combustion processes or fire accident.

Hadamard-transform fluorescence-lifetime imaging.

We discuss a Hadamard-transform-based fluorescence-lifetime-imaging (HT-FLI) technique for fluorescence-lifetime-imaging microscopy (FLIM). The HT-FLI uses a Fourier-transform phase-modulation fluorometer (FT-PMF) for fluorescence-lifetime measurements, where the modulation frequency of the excitation light is swept linearly in frequency from zero to a specific maximum during a fixed duration of time. Thereafter, fluorescence lifetimes are derived through Fourier transforms for the fluorescence and reference waveforms. The FT-PMF enables the analysis of multi-component samples simultaneously. HT imaging uses electronic exchange of HT illumination mask patterns, and a high-speed, high-sensitivity photomultiplier, to eliminate frame-rate issues that accompany two-dimensional image detectors.

Adaptive sampling dual terahertz comb spectroscopy using dual free-running femtosecond lasers.

Terahertz (THz) dual comb spectroscopy (DCS) is a promising method for high-accuracy, high-resolution, broadband THz spectroscopy because the mode-resolved THz comb spectrum includes both broadband THz radiation and narrow-line CW-THz radiation characteristics. In addition, all frequency modes of a THz comb can be phase-locked to a microwave frequency standard, providing excellent traceability. However, the need for stabilization of dual femtosecond lasers has often hindered its wide use. To overcome this limitation, here we have demonstrated adaptive-sampling THz-DCS, allowing the use of free-running femtosecond lasers. To correct the fluctuation of the time and frequency scales caused by the laser timing jitter, an adaptive sampling clock is generated by dual THz-comb-referenced spectrum analysers and is used for a timing clock signal in a data acquisition board. The results not only indicated the successful implementation of THz-DCS with free-running lasers but also showed that this configuration outperforms standard THz-DCS with stabilized lasers due to the slight jitter remained in the stabilized lasers.

Real-time absolute frequency measurement of continuous-wave terahertz radiation based on dual terahertz combs of photocarriers with different frequency spacings.

Real-time measurement of the absolute frequency of continuous-wave terahertz (CW-THz) radiation is required for characterization and frequency calibration of practical CW-THz sources. We proposed a method for real-time monitoring of the absolute frequency of CW-THz radiation involving temporally parallel, i.e., simultaneous, measurement of two pairs of beat frequencies and laser repetition frequencies based on dual THz combs of photocarriers (PC-THz combs) with different frequency spacings. To demonstrate the method, THz-comb-referenced spectrum analyzers were constructed with a dual configuration based on dual femtosecond lasers. Regardless of the presence or absence of frequency control in the PC-THz combs, a frequency precision of 10(-11) was achieved at a measurement rate of 100 Hz. Furthermore, large fluctuation of the CW-THz frequencies, crossing several modes of the PC-THz combs, was correctly monitored in real time. The proposed method will be a powerful tool for the research and development of practical CW-THz sources, and other applications.

An inducible recA expression Bacillus subtilis genome vector for stable manipulation of large DNA fragments.

BACKGROUND: The Bacillus subtilis genome (BGM) vector is a novel cloning system based on the natural competence that enables B. subtilis to import extracellular DNA fragments into the cell and incorporate the recombinogenic DNA into the genome vector by homologous recombination. The BGM vector system has several attractive properties, such as a megabase cloning capacity, stable propagation of cloned DNA inserts, and various modification strategies using RecA-mediated homologous recombination. However, the endogenous RecA activity may cause undesirable recombination, as has been observed in yeast artificial chromosome systems. In this study, we developed a novel BGM vector system of an inducible recA expression BGM vector (iREX), in which the expression of recA can be controlled by xylose in the medium. RESULTS: We constructed the iREX system by introducing the xylose-inducible recA expression cassette followed by the targeted deletion of the endogenous recA. Western blot analysis showed that the expression of recA was strictly controlled by xylose in the medium. In the absence of xylose, recA was not expressed in the iREX, and the RecA-mediated recombination reactions were greatly suppressed. By contrast, the addition of xylose successfully induced RecA expression, which enabled the iREX to exploit the same capacities of transformation and gene modifications observed with the conventional BGM vector. In addition, an evaluation of the stability of the cloned DNA insert demonstrated that the DNA fragments containing homologous sequences were more stably maintained in the iREX by suppressing undesirable homologous recombination. CONCLUSIONS: We developed a novel BGM vector with inducible recA expression system, iREX, which enables us to manipulate large DNA fragments more stably than the conventional BGM vector by suppressing undesirable recombination. In addition, we demonstrate that the iREX can be applied to handling the DNA, which has several homologous sequences, such as multiple-reporter expression cassettes. Thus, the iREX expands the utility of the BGM vector as a platform for engineering large DNA fragments.

Double-modulation reflection-type terahertz ellipsometer for measuring the thickness of a thin paint coating.

We constructed a double-modulation, reflection-type terahertz (THz) ellipsometer for precise measurement of the thickness of a paint film which is coated on a metal surface and which is not transparent to visible or mid-infrared light. The double-modulation technique enabled us to directly obtain two ellipsometric parameters, Δ(ω) and Ψ(ω), as a function of angular frequency, ω, with a single measurement while reducing flicker noise due to a pump laser. The bias voltage of a photoconductive antenna (PCA) used as a THz pulse emitter was modulated at 100 kHz, and a first lock-in amplifier (LA1) was connected to the output of an electro-optic (EO) signal-sampling unit. In addition, a wire-grid polarizer (WGP) was rotated at 100 Hz to conduct polarization modulation with a frequency of 200 Hz. The output signal from LA1 was fed into a second lock-in amplifier (LA2) that worked in synchronization with the rotating WGP (RWGP). By operating LA2 in a quadrature phase-detection mode, we were able to obtain in-phase and out-of-phase signals simultaneously, from which the two ellipsometric parameters for an isotropic sample could be derived at the same time while cancelling common-mode noise. The lower detection limit of the thickness measurement and the relative standard deviation (RSD) of a black paint film coated on an aluminum substrate were 4.3 µm and 1.4%, respectively. The possibility of determining all elements of the Jones matrix for an anisotropic material is also discussed.

Bacillus subtilis genome vector-based complete manipulation and reconstruction of genomic DNA for mouse transgenesis.

BACKGROUND: The Bacillus subtilis genome (BGM) vector is a novel cloning system for large DNA fragments, in which the entire 4.2 Mb genome of B. subtilis functions as a vector. The BGM vector system has several attractive properties, such as a large cloning capacity of over 3 Mb, stable propagation of cloned DNA and various modification strategies using RecA-mediated homologous recombination. However, genetic modifications using the BGM vector system have not been fully established, and this system has not been applied to transgenesis. In this study, we developed important additions to the genetic modification methods of the BGM vector system. To explore the potential of the BGM vector, we focused on the fish-like odorant receptor (class I OR) gene family, which consists of 158 genes and forms a single gene cluster. Although a cis-acting locus control region is expected to regulate transcription, this has not yet been determined experimentally. RESULTS: Using two contiguous bacterial artificial chromosome clones containing several class I OR genes, we constructed two transgenes in the BGM vector by inserting a reporter gene cassette into one class I OR gene. Because they were oriented in opposite directions, we performed an inversion modification to align their orientation and then fused them to enlarge the genomic structure. DNA sequencing revealed that no mutations occurred during gene manipulations with the BGM vector. We further demonstrated that the modified, reconstructed genomic DNA fragments could be used to generate transgenic mice. Transgenic mice carrying the enlarged transgene recapitulated the expression and axonal projection patterns of the target class I OR gene in the main olfactory system. CONCLUSION: We offer a complete genetic modification method for the BGM vector system, including insertion, deletion, inversion and fusion, to engineer genomic DNA fragments without any trace of modifications. In addition, we demonstrate that this system can be used for mouse transgenesis. Thus, the BGM vector system can be an alternative platform for engineering large DNA fragments in addition to conventional systems such as bacterial and yeast artificial chromosomes. Using this system, we provide the first experimental evidence of a cis-acting element for a class I OR gene.