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  1. Article ; Online: A Simple and Fast Manual Centrifuge to Spin Solutions in 96-Well PCR Plates.

    Motohashi, Ken

    Methods and protocols

    2020  Volume 3, Issue 2

    Abstract: A simple and fast manual centrifuge was developed to spin down solutions in 96-well polymerase chain reaction (PCR) plates. A commercially available salad spinner was utilized for this purpose. Acceleration and deceleration of the centrifuge were faster ... ...

    Abstract A simple and fast manual centrifuge was developed to spin down solutions in 96-well polymerase chain reaction (PCR) plates. A commercially available salad spinner was utilized for this purpose. Acceleration and deceleration of the centrifuge were faster than those of a conventional electric centrifuge using 96-well PCR plates. Solutions in a 96-well PCR plate settled quickly after centrifuging for only 3 s. This lightweight centrifuge can be stored under a laboratory bench or on a shelf and can be put on the bench only when required, whereas the electric centrifuge is immobile due to its weight and the requirement of electric cables. This simple centrifuge is inexpensive, requires minimal effort for making, and can be used anywhere.
    Language English
    Publishing date 2020-05-25
    Publishing country Switzerland
    Document type Journal Article
    ISSN 2409-9279
    ISSN (online) 2409-9279
    DOI 10.3390/mps3020041
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  2. Article ; Online: A Simple and Fast Manual Centrifuge to Spin Solutions in 96-Well PCR Plates

    Ken Motohashi

    Methods and Protocols, Vol 3, Iss 41, p

    2020  Volume 41

    Abstract: A simple and fast manual centrifuge was developed to spin down solutions in 96-well polymerase chain reaction (PCR) plates. A commercially available salad spinner was utilized for this purpose. Acceleration and deceleration of the centrifuge were faster ... ...

    Abstract A simple and fast manual centrifuge was developed to spin down solutions in 96-well polymerase chain reaction (PCR) plates. A commercially available salad spinner was utilized for this purpose. Acceleration and deceleration of the centrifuge were faster than those of a conventional electric centrifuge using 96-well PCR plates. Solutions in a 96-well PCR plate settled quickly after centrifuging for only 3 s. This lightweight centrifuge can be stored under a laboratory bench or on a shelf and can be put on the bench only when required, whereas the electric centrifuge is immobile due to its weight and the requirement of electric cables. This simple centrifuge is inexpensive, requires minimal effort for making, and can be used anywhere.
    Keywords centrifuge ; 0.2-mL 96-well PCR plate ; high-throughput screening ; PCR solution ; spin-down ; Biology (General) ; QH301-705.5
    Language English
    Publishing date 2020-05-01T00:00:00Z
    Publisher MDPI AG
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  3. Article ; Online: A novel series of high-efficiency vectors for TA cloning and blunt-end cloning of PCR products.

    Motohashi, Ken

    Scientific reports

    2019  Volume 9, Issue 1, Page(s) 6417

    Abstract: An efficient PCR cloning method is indispensable in modern molecular biology, as it can greatly improve the efficiency of DNA cloning processes. Here, I describe the development of three vectors for TA cloning and blunt-end cloning. Specifically, pCRT ... ...

    Abstract An efficient PCR cloning method is indispensable in modern molecular biology, as it can greatly improve the efficiency of DNA cloning processes. Here, I describe the development of three vectors for TA cloning and blunt-end cloning. Specifically, pCRT and pCRZeroT were designed to improve the efficiency of TA cloning. pCRZeroT can also be used with pCRZero to facilitate blunt-end cloning using the ccdB gene. Using pCRZero and pCRZeroT and applying the Golden Gate reaction, I developed a direct PCR cloning protocol with non-digested circular vectors and PCR products. This direct PCR cloning protocol yielded colony-formation rates and cloning efficiencies that are comparable with those obtained by conventional PCR cloning with pre-digested vectors and PCR products. The three plasmids I designed are available from Addgene ( https://www.addgene.org/ ).
    MeSH term(s) Bacterial Proteins/genetics ; Bacterial Proteins/metabolism ; Base Sequence ; Cloning, Molecular/methods ; DNA Primers/chemistry ; DNA Primers/metabolism ; Escherichia coli/genetics ; Escherichia coli/metabolism ; Gene Expression ; Genetic Engineering/methods ; Humans ; Plasmids/chemistry ; Plasmids/metabolism ; Polymerase Chain Reaction/methods
    Chemical Substances Bacterial Proteins ; CcdB protein, Plasmid F ; DNA Primers
    Language English
    Publishing date 2019-04-23
    Publishing country England
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ZDB-ID 2615211-3
    ISSN 2045-2322 ; 2045-2322
    ISSN (online) 2045-2322
    ISSN 2045-2322
    DOI 10.1038/s41598-019-42868-6
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  4. Article ; Online: Development of highly sensitive and low-cost DNA agarose gel electrophoresis detection systems, and evaluation of non-mutagenic and loading dye-type DNA-staining reagents.

    Motohashi, Ken

    PloS one

    2019  Volume 14, Issue 9, Page(s) e0222209

    Abstract: Highly sensitive and low-cost DNA agarose gel detection systems were developed using non-mutagenic and loading dye-type DNA-staining reagents. The DNA detection system that used Midori Green Direct and Safelook Load-Green, both with an optimum excitation ...

    Abstract Highly sensitive and low-cost DNA agarose gel detection systems were developed using non-mutagenic and loading dye-type DNA-staining reagents. The DNA detection system that used Midori Green Direct and Safelook Load-Green, both with an optimum excitation wavelength at ~490 nm, could detect DNA-fragments at the same sensitivity to that of the UV (312 nm)-transilluminator system combined with ethidium bromide, after it was excited by a combination of cyan LED light and a shortpass filter (510 nm). The cyan LED system can be also applied to SYBR Safe that is widely used as a non-toxic dye for post-DNA-staining. Another DNA-detection system excited by black light was also developed. Black light used in this system had a peak emission at 360 nm and caused less damage to DNA due to lower energy of UV rays with longer wavelength when compared to those of short UV rays. Moreover, hardware costs of the black light system were ~$100, less than 1/10 of the commercially available UV (365 nm) transilluminator (>$1,000). EZ-Vision and Safelook Load-White can be used as non-mutagenic and loading dye-type DNA-staining reagents in this system. The black light system had a greater detection sensitivity for DNA fragments stained by EZ-Vision and Safelook Load-White compared with the commercially available imaging system using UV (365 nm) transilluminator.
    MeSH term(s) DNA/analysis ; Electrophoresis, Agar Gel/economics ; Electrophoresis, Agar Gel/methods ; Ethidium ; Staining and Labeling
    Chemical Substances DNA (9007-49-2) ; Ethidium (EN464416SI)
    Language English
    Publishing date 2019-09-09
    Publishing country United States
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ISSN 1932-6203
    ISSN (online) 1932-6203
    DOI 10.1371/journal.pone.0222209
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  5. Article ; Online: A novel series of high-efficiency vectors for TA cloning and blunt-end cloning of PCR products

    Ken Motohashi

    Scientific Reports, Vol 9, Iss 1, Pp 1-

    2019  Volume 11

    Abstract: Abstract An efficient PCR cloning method is indispensable in modern molecular biology, as it can greatly improve the efficiency of DNA cloning processes. Here, I describe the development of three vectors for TA cloning and blunt-end cloning. Specifically, ...

    Abstract Abstract An efficient PCR cloning method is indispensable in modern molecular biology, as it can greatly improve the efficiency of DNA cloning processes. Here, I describe the development of three vectors for TA cloning and blunt-end cloning. Specifically, pCRT and pCRZeroT were designed to improve the efficiency of TA cloning. pCRZeroT can also be used with pCRZero to facilitate blunt-end cloning using the ccdB gene. Using pCRZero and pCRZeroT and applying the Golden Gate reaction, I developed a direct PCR cloning protocol with non-digested circular vectors and PCR products. This direct PCR cloning protocol yielded colony-formation rates and cloning efficiencies that are comparable with those obtained by conventional PCR cloning with pre-digested vectors and PCR products. The three plasmids I designed are available from Addgene (https://www.addgene.org/).
    Keywords Medicine ; R ; Science ; Q
    Language English
    Publishing date 2019-04-01T00:00:00Z
    Publisher Nature Publishing Group
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  6. Article ; Online: Development of highly sensitive and low-cost DNA agarose gel electrophoresis detection systems, and evaluation of non-mutagenic and loading dye-type DNA-staining reagents.

    Ken Motohashi

    PLoS ONE, Vol 14, Iss 9, p e

    2019  Volume 0222209

    Abstract: Highly sensitive and low-cost DNA agarose gel detection systems were developed using non-mutagenic and loading dye-type DNA-staining reagents. The DNA detection system that used Midori Green Direct and Safelook Load-Green, both with an optimum excitation ...

    Abstract Highly sensitive and low-cost DNA agarose gel detection systems were developed using non-mutagenic and loading dye-type DNA-staining reagents. The DNA detection system that used Midori Green Direct and Safelook Load-Green, both with an optimum excitation wavelength at ~490 nm, could detect DNA-fragments at the same sensitivity to that of the UV (312 nm)-transilluminator system combined with ethidium bromide, after it was excited by a combination of cyan LED light and a shortpass filter (510 nm). The cyan LED system can be also applied to SYBR Safe that is widely used as a non-toxic dye for post-DNA-staining. Another DNA-detection system excited by black light was also developed. Black light used in this system had a peak emission at 360 nm and caused less damage to DNA due to lower energy of UV rays with longer wavelength when compared to those of short UV rays. Moreover, hardware costs of the black light system were ~$100, less than 1/10 of the commercially available UV (365 nm) transilluminator (>$1,000). EZ-Vision and Safelook Load-White can be used as non-mutagenic and loading dye-type DNA-staining reagents in this system. The black light system had a greater detection sensitivity for DNA fragments stained by EZ-Vision and Safelook Load-White compared with the commercially available imaging system using UV (365 nm) transilluminator.
    Keywords Medicine ; R ; Science ; Q
    Subject code 612
    Language English
    Publishing date 2019-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  7. Article: [MASS EXAMINATION OF VISUALLY IMPAIRED SUBJECTS IN SHIZUOKA-KEN].

    MOTOHASHI, T

    Rinsho ganka. Japanese journal of clinical ophthalmology

    1964  Volume 18, Page(s) 1051–1057

    MeSH term(s) Adolescent ; Blindness ; Child ; Eye Diseases ; Geriatrics ; Humans ; Infant ; Japan ; Mass Screening ; Statistics as Topic ; Vision Tests
    Language Japanese
    Publishing date 1964-09
    Publishing country Japan
    Document type Journal Article
    ZDB-ID 800088-8
    ISSN 0370-5579
    ISSN 0370-5579
    Database MEDical Literature Analysis and Retrieval System OnLINE

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    In stock of ZB MED Cologne/Königswinter

    Zs.B 136: Show issues Location:
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    bis Jg. 2021: Bestellungen von Artikeln über das Online-Bestellformular
    ab Jg. 2022: Lesesaal (EG)

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  8. Article: Evaluation of the efficiency and utility of recombinant enzyme-free seamless DNA cloning methods.

    Motohashi, Ken

    Biochemistry and biophysics reports

    2017  Volume 9, Page(s) 310–315

    Abstract: Simple and low-cost recombinant enzyme-free seamless DNA cloning methods have recently become available. ...

    Abstract Simple and low-cost recombinant enzyme-free seamless DNA cloning methods have recently become available.
    Language English
    Publishing date 2017-01-26
    Publishing country Netherlands
    Document type Journal Article
    ZDB-ID 2831046-9
    ISSN 2405-5808
    ISSN 2405-5808
    DOI 10.1016/j.bbrep.2017.01.010
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  9. Article ; Online: Seamless Ligation Cloning Extract (SLiCE) Method Using Cell Lysates from Laboratory Escherichia coli Strains and its Application to SLiP Site-Directed Mutagenesis.

    Motohashi, Ken

    Methods in molecular biology (Clifton, N.J.)

    2017  Volume 1498, Page(s) 349–357

    Abstract: Cell lysates from laboratory Escherichia coli strains endogenously exhibit homologous recombination activity, which can be utilized for seamless DNA cloning in vitro. This method, termed Seamless Ligation Cloning Extract (SLiCE) cloning, enables high ... ...

    Abstract Cell lysates from laboratory Escherichia coli strains endogenously exhibit homologous recombination activity, which can be utilized for seamless DNA cloning in vitro. This method, termed Seamless Ligation Cloning Extract (SLiCE) cloning, enables high cloning efficiency with simultaneous integration of two unpurified DNA fragments into a vector. In addition, the SLiCE method is highly cost-effective, as several laboratory E. coli strains may be utilized as sources of SLiCE. Previously, the SLiCE technique has been applied to site-directed mutagenesis to develop a novel technique termed SLiCE-mediated polymerase chain reaction (PCR)-based site-directed mutagenesis (SLiP site-directed mutagenesis). Two DNA fragments containing a mutation site can be simultaneously integrated into a vector while avoiding the introduction of undesirable mutations in the vector. Therefore, SLiP site-directed mutagenesis simplifies multiple procedures involved in PCR-based site-directed mutagenesis such as overlap extension method PCR or the Megaprimer method.
    Language English
    Publishing date 2017
    Publishing country United States
    Document type Journal Article
    ISSN 1940-6029
    ISSN (online) 1940-6029
    DOI 10.1007/978-1-4939-6472-7_23
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  10. Article ; Online: Evaluation of the efficiency and utility of recombinant enzyme-free seamless DNA cloning methods

    Ken Motohashi

    Biochemistry and Biophysics Reports, Vol 9, Iss C, Pp 310-

    2017  Volume 315

    Abstract: Simple and low-cost recombinant enzyme-free seamless DNA cloning methods have recently become available. In vivo Escherichia coli cloning (iVEC) can directly transform a mixture of insert and vector DNA fragments into E. coli, which are ligated by ... ...

    Abstract Simple and low-cost recombinant enzyme-free seamless DNA cloning methods have recently become available. In vivo Escherichia coli cloning (iVEC) can directly transform a mixture of insert and vector DNA fragments into E. coli, which are ligated by endogenous homologous recombination activity in the cells. Seamless ligation cloning extract (SLiCE) cloning uses the endogenous recombination activity of E. coli cellular extracts in vitro to ligate insert and vector DNA fragments. An evaluation of the efficiency and utility of these methods is important in deciding the adoption of a seamless cloning method as a useful tool. In this study, both seamless cloning methods incorporated inserting DNA fragments into linearized DNA vectors through short (15–39 bp) end homology regions. However, colony formation was 30–60-fold higher with SLiCE cloning in end homology regions between 15 and 29 bp than with the iVEC method using DH5α competent cells. E. coli AQ3625 strains, which harbor a sbcA gene mutation that activates the RecE homologous recombination pathway, can be used to efficiently ligate insert and vector DNA fragments with short-end homology regions in vivo. Using AQ3625 competent cells in the iVEC method improved the rate of colony formation, but the efficiency and accuracy of SLiCE cloning were still higher. In addition, the efficiency of seamless cloning methods depends on the intrinsic competency of E. coli cells. The competency of chemically competent AQ3625 cells was lower than that of competent DH5α cells, in all cases of chemically competent cell preparations using the three different methods. Moreover, SLiCE cloning permits the use of both homemade and commercially available competent cells because it can use general E. coli recA− strains such as DH5α as host cells for transformation. Therefore, between the two methods, SLiCE cloning provides both higher efficiency and better utility than the iVEC method for seamless DNA plasmid engineering.
    Keywords Homologous recombination ; in vivo Escherichia coli cloning ; Seamless DNA cloning ; SLiCE ; Biology (General) ; QH301-705.5 ; Biochemistry ; QD415-436
    Subject code 630
    Language English
    Publishing date 2017-03-01T00:00:00Z
    Publisher Elsevier
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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