Gene Engineering Division

Research tools

• Artificial enzymatic RNA editing system
  • RNA editing of BFP, a point mutant of GFP, using artificial APOBEC1 deaminase to restore the genetic code.
    Bhakta S, Sakari M, Tsukahara T.
    Sci. Rep. 10 (1): 17304 (2020). PMID 33057101.

    Catalog no.	Name of resource	Descriptiion
    RDB19359	Target BFP gene expression plasmid	Expression vector of target BFP gene, a point mutant (T199C) of GFP for artificial enzymatic RNA editing system.
    RDB19360	pCS2+MT-MS2HB-APOBEC1	Expression vector of the catalytic domain of human APOBEC1 tagged with MS2HB at N-terminus for artificial enzymatic RNA editing system.
    RDB19361	MS2-guideRNA21bp	Expression vector of guideRNA, a 21 ntd sequence complementary to the target BFP mRNA tagged with MS2 stem loop at C-terminus for artificial enzymatic RNA editing system.

• Auxin Inducible Degron (AID) System
  • Using the AID System, you can degrade proteins fused with a degron (a small domain that can induce protein degradation) in a very short period of time.
• ChIP-seq analysis
  • ChIP-seq analysis of genomic binding regions of five major transcription factors highlights a central role for ZIC2 in the mouse epiblast stem cell gene regulatory network.
    Matsuda, K., Mikami, T., Oki, S., Iida, H., Andrabi, M., Boss, J.M., Yamaguchi, K., Shigenobu, S., Kondoh, H.
    Development 144 (11): 1948-1958 (2017). PubMed PMID 28455373.

    Catalog no.	Name of resource	Descriptiion
    RDB15774	pCAGGS-BLRP-BirA	Expression vector to perform ChIP-seq analysis using biotinylated proteins.

    ChIP-seq is a powerful method of comprehensive analysis of transcription factor (TF) binding sites. However, the bottle neck of ChIP-seq analysis is the quality of antibody, and thus many researchers have been given up. To overcome, authors constructed the vector expressing “biotinylated-tag” transcription factors. With this vector, you can get acceptable level of TF-DNA complex for ChIP-seq analysis by streptavidin-beads for any kinds of TF.

• Cre-loxP and FLP-FRT system
• Host strain
  • Host strain for producing recombinant proteins incorporating synthetic amino acids
  • Host strain for selective isotope labeling
  • Bifidobacterium longum 105-A
  • Bacillus stearothermophilus K1041
• MAPK p38 inhibitor screening by using E. coli.
• Open-Sandwich Immunoassay (OS-IA)
  • Homogeneous Noncompetitive Luminescent Immunodetection of Small Molecules by Ternary Protein Fragment Complementation.
    Ohmuro-Matsuyama, Y., Ueda, H.
    Anal. Chem. 90 (5): 3001-3004 (2018). PubMed PMID 29446920.

    Catalog no.	Name of resource	Descriptiion
    RDB16003	pET-LnBiT	Bacterial expression vector of LnBiT, a comporntnt of the Open Sandwich Bioluminescent Immunoassay (OS-BLIA).

• Optogenetics resource
  • hannelrhodopsin-green receiver (ChRGR)
  • Chimera Na+-Pump Rhodopsin
  • Lanthanide nanoparticle-channelrhodopsin system
  • C2C12 cells stably expressing ChR2 (ChR2-C2C12)
  • Mouse strain expressing channelrhodopsin
• Thermostable kanamycin for microbial gene disruption
• Tet inducible system

Genome Editing

• Genome Editing by CRISPR/Cas9
  • Expression vector of Cas9 enzyme
  • Plasmids for the evaluation of the efficiency of genome editing
  • Knock-in markers with CRISPR/Cas9 genome editing
  

Fluorescent & Luminescent proteins

Fluorescent protein resource

• Cyan
  • Kusabira Cyan
  • Midoriishi-Cyan (MiCy, mMiCy1)
• Green
  • Umikinoko Green (mUKG1, hmUKG1)
  • Azami Green (AG, hmAG1, hmAG1, hmAG407)
  • h2-3
  • GFP (GFPS1)
  • StayGold (StayGold, oxStayGold, tdStayGold, tdoxStayGold)
• Yellow
  • Achilles
  • Venus (Venus, mVenus, cp49Venus, cp145Venus, cp157Venus, cp173Venus, cp195Venus, cp229Venus)
  • Cy11.5
• Orange
  • Kusabira Orange (KO1, hKO1, mKO1, hmKO1, mKO2, hmKO2, hmKO-K)
• Red
  • AzaleaB5
  • Keima570 and Keima-red (dKeima570, hdKeima570, dKeima-Red, mKeima-Red, hdKeima-Red, hmKeima-Red, tdKeima)
  • NpF2164g5_BV4
• Photoconversion type
  • Kaede
  • Kikume Green-Red (KikGR, mKikGR, mKikGR13.2, Xpa)
• Photochromism type
  • Dronpa-Green (Dronpa, Dronpa2, Dronpa3, 22G)
• Timer type
  • Kusabira Green Orange (mK-GO)
• Destabilized- and Secreted-FPs
• Non-fluorescent chromoprotein
  • cjBlue (cjBlue, cjBlue Y64L)
• Sea cactus therostable GFP

Higher intensity luciferases having Green-, Yellow- or Red-emission by using D-luciferin

• These luciferases have at most four-time maximum luminous intensity than that of widely used luciferase of the Photinus pyralis, a common North American firefly. Further more, Green-, Yellow- or Red-emission can be obtained by using D-luciferin as a substrate of each luciferase.

AkaLuc luciferase providing brighter and red-shifted luminescence

• The artificial bioluminescence system AkaBLI enables noninvasive signal observation in deep tissue of living animals. It was developed by Dr. Atsushi Miyawaki and Dr. Satoshi Iwano of the RIKEN Center for Brain Science, and Dr. Shojiro Maki of the University of Electro-Communications. The AkaBLI consists of an artificial substrate AkaLumine with improved tissue permeability and an artificial luciferase Akaluc optimized to AkaLumine. The intensity of the luminescence of AkaBLI system is 100 to 1000 folds brighter than the conventional systems.

Nano-lantern luminescent and fluorescent protein

• Nano lantern (NL) consists of Renilla luciferase and adjacent fluorescence protein. The chemiluminescence of the luciferase provides light source for excitation and enables the fluorescence protein to be observed. Three colors of NLs, yellow, cyan and orange, have been developed. NLs do not require external light source and overcome problems such as autofluorescence, phototoxicity, and photobleaching.

Sensor & Visualization

• Sensor & Visualization
  • Bilirubin indicator UnaG, BReleaCa
  • cAMP indicator
  • Caspase activity indicator
  • pH sensor
  • Protein concentration in cells
  • RA indicator
  • Voltage indicator
• ATP measurement
  • QUEEN
    Diversity in ATP concentrations in a single bacterial cell population revealed by quantitative single-cell imaging.
    Yaginuma, H., Kawai, S., Tabata, K.V., Tomiyama, K., Kakizuka, A., Komatsuzaki, T., Noji, H., Imamura, H.
    Sci. Rep. 4: 6522 (2014). PMID 25283467.
• Autophagy and Mitophagy indicator
  • Chaperone mediated autophagy activity by the GAPDH-HT indicator by Dr. Takahiro Seki’s lab as well as fluorescent protein probes of LC3B accumulation developed by Dr. Noboru Mizushima’s lab, Dr. Itaru Hamachi’s lab and Dr. Keiji Kimura’s lab are available.
• Calcium-ion sensor
  • Fluorescent protein-based Ca²⁺ sensors, which are composed with calmodulin, fluorescent protein and M13 peptide (CaM binding domain of myosin light chain kinase), are designated to visualize intracellular [Ca²⁺] dynamics.
  • Various improvements of the Ca²⁺ sensors have been made for optimum temperature, signal/noise ratio, susceptibility and so on.
  • Many papers using Ca²⁺ sensors have been published reporting the estimation of calcium concentration.
  • G-CaMP
  • Pericam
  • Yellow Cameleon
• Cell cycle indicator Fucci
  • To monitor cell cycle progression in living cells, cell cycle indicator Fucci probes deveoped by Dr. Atsushi Miyawaki’s lab are available.
• Epigenetics reporter
  • Visualization of histone acetylation: The Histac fluorescent probes deposited by Dr. Kazuki Sasaki allow you monitoring the state of activity of acetylation of histone H4 by fluorescence in living cells.
  • Visualization of methylated DNA: The EGFP-MBD-nls protein recognizes the methylated DNA and you can follow status of the DNA methylation in situ under physiological conditions using the pEGFP-MBD-nls expression clone.
• Knock in markers with CRISPR/Cas9 genome editing
  • As a part of Auxin Inducible Degron (AID) System clones, Dr. Masato Kanemaki provides a series of knock-in fluorescent and selection markers with CRISPR/Cas9 genome editing.
• Notch signaling reporter
  • The pRBS-EGFP and RBP-J-Venus expression clones deposited by Dr. Makoto Mark Taketo and Dr. Kenji Tanigaki, respectively, allow you monitoring the state of activation of the Notch signaling by fluorescence in living cells.
• Visualization of Organelles
  • Organelle marker/subcellular localization: We are providing genetic resources for visualization of organelles such as mitochondria and nucleus. Each clone contains an organelle localization signal sequence fused with fluorescent proteins or epitope tags. Organelles can be detected by fluorescence or by detecting epitope tags with antibodies.
  • Visualizing multiple inter-organelle contact sites: Divided GFP-fragment have a feature that they emit fluorescence again by reconstitution when each fragment is in close proximity. When labelled organelles such as mitochondria and ERs by the divided GFPs exist together, the contact sites in cells are visualized by the reconstituted fluorescent protein. The vectors of fluorescent protein probes were developed by Dr. Yasushi Tamura of the Yamagata University, Dr. Toshiya Endo of the Kyoto Sangyo University.
    
  • GPI anchored Reconstitution-Activated Proteins Highlight Intercellular Connections (GRAPHIC) developed by Dr. Natatoki Kinoshita.
• Sphingolipid marker
  • Lipid rafts are small lipid domains on the cell membrane and are thought to play an important role in signal transduction, endocytosis and more. We provide fluorescent probes for sphingomyelin and cholesterol lipid domains.
  • Nakanori: sphingomyelin and cholesterol lipid domain (lipid raft)
  • D4 toxin: cholesterol rich domain
  • lysenin: sphingomyelin

Mammalian cell

• Promoter collection
  • Promoter collection
  • Fire fly luciferase (assayed)
  • Renilla luciferase (assayed)
• Control vector
  • Luciferase, lacZ
• Recombinant virus hub
• Backbone vector

E. coli

• Backbone vector

Thermus thermophilus HB8

• Gene disruption plasmid

Schizosaccharomyces pombe

• S. pombe ORFeome clone
• Expression Vector Backbone of Schizosaccharomyces pombe

Saccharomyces cerevisiae

• Backbone vector

Other resources

• Gene set collection
• Biomass collection

(2006.02.16 T.M.)

2022.05.13