The CRISPR/Cas9 system provides a powerful tool for genome editing in various model organisms, including zebrafish. The establishment of targeted gene-disrupted zebrafish (knockouts) is readily ...achieved by CRISPR/Cas9-mediated genome modification. Recently, exogenous DNA integration into the zebrafish genome via homology-independent DNA repair was reported, but this integration contained various mutations at the junctions of genomic and integrated DNA. Thus, precise genome modification into targeted genomic loci remains to be achieved. Here, we describe efficient, precise CRISPR/Cas9-mediated integration using a donor vector harbouring short homologous sequences (10-40 bp) flanking the genomic target locus. We succeeded in integrating with high efficiency an exogenous mCherry or eGFP gene into targeted genes (tyrosinase and krtt1c19e) in frame. We found the precise in-frame integration of exogenous DNA without backbone vector sequences when Cas9 cleavage sites were introduced at both sides of the left homology arm, the eGFP sequence and the right homology arm. Furthermore, we confirmed that this precise genome modification was heritable. This simple method enables precise targeted gene knock-in in zebrafish.
Genome editing-assisted gene knock-in technology has become rather diversifieda nd improved from day to day. Since genome editing relies on the endogenous DSB repair machineries, their appropriate ...utilization is a key to control the editing outcome.
Endogenous retroviruses (ERVs) are genomic elements derived from retroviral infections in ancestral germ lines. Most ERVs are inactivated by genetic or epigenetic mechanisms, such as DNA methylation. ...RD-114-virus-related sequence (RDRS) C2a is a feline endogenous retrovirus present in all domestic cats; however, its expression and function are not clearly known. DNA methylation at CpG dinucleotides is a hallmark of silenced ERVs. This study aimed to investigate whether long terminal repeats (LTRs) of RDRS C2a function as a gene regulatory region. The DNA methylation status of RDRS C2a was examined by bisulfite sequencing, and CpG sites in 5ʹ LTR of RDRS C2a were found hypomethylated, whereas those in 3ʹ LTR were hypermethylated in feline cells. Several transcription factor-binding sites were identified in LTRs of RDRS C2a. Luciferase assay suggested that 5ʹ LTR of RDRS C2a exhibited strong transcriptional activity, which was suppressed by in vitro DNA methylation. The study indicates that 5ʹ LTR of RDRS C2a possibly functions as a promoter for itself or neighboring genes.
Although near-infrared photons in telecommunication bands are required for long-distance quantum communication, various quantum information tasks have been performed by using visible photons for the ...past two decades. Recently, such visible photons from diverse media including atomic quantum memories have also been studied. Optical frequency down-conversion from visible to telecommunication bands while keeping the quantum states is thus required for bridging such wavelength gaps. Here we report demonstration of a quantum interface of frequency down-conversion from visible to telecommunication bands by using a nonlinear crystal, which has a potential to work over wide bandwidths, leading to a high-speed interface of frequency conversion. We achieved the conversion of a picosecond visible photon at 780 nm to a 1,522-nm photon, and observed that the conversion process retained entanglement between the down-converted photon and another photon.
The segmental pattern of somites is generated by sequential conversion of the temporal periodicity provided by the molecular clock. Whereas the basic structure of this clock is conserved among ...different species, diversity also exists, especially in terms of the molecular network. The temporal periodicity is subsequently converted into the spatial pattern of somites, and Mesp2 plays crucial roles in this conversion in the mouse. However, it remains unclear whether Mesp genes play similar roles in other vertebrates. In this study, we generated zebrafish mutants lacking all four zebrafish Mesp genes by using TALEN-mediated genome editing. Contrary to the situation in the mouse Mesp2 mutant, in the zebrafish Mesp quadruple mutant embryos the positions of somite boundaries were clearly determined and morphological boundaries were formed, although their formation was not completely normal. However, each somite was caudalized in a similar manner to the mouse Mesp2 mutant, and the superficial horizontal myoseptum and lateral line primordia were not properly formed in the quadruple mutants. These results clarify the conserved and species-specific roles of Mesp in the link between the molecular clock and somite morphogenesis.
Methods to prevent collisions between people to avoid traffic accidents are receiving significant attention. To measure the position in the non-line-of-sight (NLOS) area, which cannot be directly ...visually recognized, position-measuring methods use wireless-communication-type GPS and propagation characteristics of radio signals, such as received signal strength indication (RSSI). However, conventional position estimation methods using RSSI require multiple receivers, which decreases the position estimation accuracy, owing to the presence of surrounding buildings. This study proposes a system to solve this challenge using a receiver and position estimation method based on RSSI MAP simulation and particle filter. Moreover, this study utilizes BLE peripheral/central functions capable of advertising as the transmitter/receiver. By using the advertising radio waves, our method provides a framework for estimating the position of unspecified transmitters. The effectiveness of the proposed system is evaluated in this study through simulations and experiments in actual environments. We obtained an error average of the distance to be 1.6 m from the simulations, which shows the precision of the proposed method. In the actual environment, the proposed method showed an error average of the distance to be 3.3 m. Furthermore, we evaluated the accuracy of the proposed method when both the transmitter and receiver are in motion, which can be considered as a moving person in the outdoor NLOS area. The result shows an error of 4.5 m. Consequently, we concluded that the accuracy was comparable when the transmitter is stationary and when it is moving. Compared with conventional path loss, the model can measure distances of 3 m to 10 m, whereas the proposed method can estimate the “position” with the same accuracy in an outdoor environment. In addition, it can be expected to be used as a collision avoidance system that confirms the presence of strangers in the NLOS area.
There has been an increasing interest in mobile manipulators that are capable of performing physical work in living spaces worldwide, corresponding to an aging population with declining birth rates ...with the expectation of improving quality of life (QoL). We assume that overall research and development will accelerate by using a common robot platform among a lot of researchers since that enables them to share their research results. Therefore we have developed a compact and safe research platform, Human Support Robot (HSR), which can be operated in an actual home environment and we have provided it to various research institutes to establish the developers community. Currently, the number of HSR users is expanding to 44 sites in 12 countries worldwide (as of November 30th, 2018). To activate the community, we assume that the robot competition will be effective. As a result of international public offering, HSR has been adopted as a standard platform for international robot competitions such as RoboCup@Home and World Robot Summit (WRS). HSR is provided to participants of those competitions. In this paper, we describe HSR’s development background since 2006, and technical detail of hardware design and software architecture. Specifically, we describe its omnidirectional mobile base using the dual-wheel caster-drive mechanism, which is the basis of HSR’s operational movement and a novel whole body motion control system. Finally, we describe the verification of autonomous task capability and the results of utilization in RoboCup@Home in order to demonstrate the effect of introducing the platform.
Sea urchins have been used as model organisms in developmental biology research and the genomes of several sea urchin species have been sequenced. Recently, genome editing technologies have become ...available for sea urchins, and methods for gene knockout using the CRISPRCas9 system have been established.
is an important marine fishery resource with edible gonads. Although
has been used as a biological research material, its genome has not yet been published, and it is a non-model sea urchin for molecular biology research. However, as recent advances in genome editing technology have facilitated genome modification in non-model organisms, we applied genome editing using the CRISPR-Cas9 system to
. In this study, we targeted genes encoding ETS transcription factor (
) and pigmentation-related polyketide synthase (
). Gene fragments were isolated using primers designed by inter-specific sequence comparisons within Echinoidea. When
gene was targeted using two sgRNAs, one successfully introduced mutations and impaired skeletogenesis. In the
gene knockout, when two sgRNAs targeting the close vicinity of the site corresponding to the target site that showed 100% mutagenesis efficiency of the
gene in
, mutagenesis was not observed. However, two other sgRNAs targeting distant sites efficiently introduced mutations. In addition,
knockout
exhibited an albino phenotype in the pluteus larvae and adult sea urchins after metamorphosis. This indicates that the CRISPRCas9 system can be used to modify the genome of the non-model sea urchin
.
The characteristics of power transfer between super-modes in coupled multicore fiber are investigated numerically and experimentally. We reveal that fiber twisting and macrobending cause mode ...coupling through an adiabatic/nonadiabatic process, which is different from the mode coupling caused by structural perturbation, namely microbending. We also investigate the impulse response characteristics of coupled two-core fibers, and find that there is an optimum core pitch for minimizing the complexity of the MIMO processing. Finally, we conduct a 4 × 4 MIMO transmission experiment using two-core fibers to evaluate their transmission characteristics. We show that strongly coupled multicore fiber can reduce the quality difference between the received signals even when the transmission line has large core-dependent gain/loss values.
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