National Institute for Basic Biology

Research Support

NIBB BIORESOURCE CENTER

HeadFUJIMORI, Toshihiko

To understand the mechanisms of living organisms, studies
focusing on each gene in the design of life (the genome)
are required. The use of model animals and plants, such as
mice, Medaka (Oryzias latipes), zebraﬁsh, Arabidopsis,
Lotus japonicus, and Physcomitrella patens, make it possible
to produce genetically controlled organisms, which havemarkers placed on them, using genetic and cell engineering
technologies. Such marking allows us to conduct detailed
studies of genes and cell functions. Because these modelorganisms mature in a short period of time, changes in cells,organs, and individuals can be thoroughly and efﬁciently
observed. On this front, the NIBB BioResource Center has
the equipment, facilities, and staff to safely, efﬁciently, andappropriately maintain such organisms.

MODEL ANIMAL RESEARCH FACILITY
Associate Professor:
Technical Staff:
Technical Assistant:

WATANABE, Eiji
OHSAWA, Sonoko
NOGUCHI, Yuji
TAKAGI, Yukari
SUGINAGA, Tomomi
FUJIMOTO, Daiji
KITAZUMI, Noriaki

The worldwide genome project has almost been completedand basic biological research is now in a post-genome erain which researchers focus on investigating the functionsof individual genes. To promote the functional analysis of agene of interest, it is essential to utilize genetically alteredmodel organisms which are generated using genetic engineering technology, and harness techniques such as genedeletion, gene replacement and point mutation.
The NIBB Center for Transgenic Animals and Plants wasestablished in April 1998 to support research using transgenic and gene targeting techniques at NIBB. The NIBB Centerfor Transgenic Animals and Plants was integrated into theNIBB BioResource center in April 2010, and was renamed“The Model Animal Research Facility”; a place where technical and supporting staff develop and promote researchsupporting activities. Furthermore, a state-of-the-art facility
for transgenic animals was also opened at the end of 2003 inthe Yamate area of NIBB.

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The activities of the model animal research facility are asfollows:
1. The provision of information, materials, techniques, andanimal housing spaces to researchers.
2. T he use of various kinds of instruments to analyze
mutant, transgenic, and gene-targeted animals.
3. The development of novel techniques related to transgenic and gene targeting technology.
4. Cryopreservation and storage of transgenic mice strains.
5. G enerating genetically-engineered mice using theCRISPR/Cas9 method.

I. Research support activities (mouse)

In 2001, the NIBB mouse facility (built under specificpathogen free (SPF) conditions) was opened in the Myodaiji
area of NIBB. Since then, the production, breeding, analysis,
cryopreservation and storage of genetically manipulatedmouse strains has been conducted there ever since. The newcenter facility building in the Yamate area has strengthened
research activities that require genetically altered organisms.
The building has five floors and a total floor space of 2,500
m2 in which we can generate, breed, store and analyze transgenic, gene targeting, and mutant mice under SPF conditions.
The mouse housing area was constructed based on a barriersystem. This building is also equipped with breeding areas
for small transgenic fish and birds.

Figure 1. Mouse breeding room in the Yamate area

Figure 2. Breeding cages for replacement in the clean corridorFrom April 1, 2021 to March 31, 2022, 3,388 mice (1 transgenic line and wild-type) were brought into the facility in theYamate area, and 30,944 mice (including pups bred in thefacility) were taken out.
A number of strains of genetically altered mice from outside
the facility were brought into this area after microbiologicalcleaning using in vitro fertilization-embryo transfer techniques (1 transgenic line), and stored using cryopreservation(34 transgenic lines). The frozen eggs of 43 mice lines weretaken out of the facility.
Genome editing experiments were performed on two kindsof target genes. We generated gRNAs of the target genes,
which were transferred into fertilized eggs with Cas9 protein,
and were able to introduce intended mutations into thegenome DNA.

medaka and zebrafish, respectively. Additionally, water canbe maintained to suit the conditions desired for fish breeding.
A medaka line that allows gene induction by heat treatment,in combination with a cre/loxP system, has been developed using this facility. All the rooms are qualified to meetthe criteria for transgenic animals, allowing researchers togenerate and maintain these important biological tools.
In 2021, 0 zebrafish (0 fertilized eggs) were brought to thefacility nor were there any fertilized eggs or chicken embryos
brought in or taken from the laboratory. The animals housed
within the facility were used for research activities in neurobiology and developmental biology.

III. Research activities

The associate professor of this center, Dr. Eiji Watanabe,
is the principal investigator of the Laboratory of
Neurophysiology, which studies various mechanisms of thevisual system using a psychophysical approach. For moredetails, please refer to the laboratory’s page.

MODEL PLANT RESEARCH FACILITY
Plant Culture Laboratory

Assistant Professor:
Technical Staff:
Technical Assistant:

HOSHINO, Atsushi
TSUGANE, Kazuo
MOROOKA, Naoki
YAMAGUCHI, Chinami

Figure 3. Large sized autoclave in the Yamate area

II. Research support activities (small fish and birds)

The first floor of the center facility building in the Yamate
area provides space and facilities to maintain small fish andchick embryos. In the laboratory room for chick embryos, alarge incubation chamber is provided and set at 37.5 degrees(suitable for chick embryogenesis). The researchers canmanipulate these embryos under optimal conditions, thus
removing biohazard risks.
For researchers who employ fish as an experimental model,
480 tanks (1 liter) and 450 tanks (3 liter) are available forFigure 4. Vinyl isolators in the quarantine room

The Plant Culture Laboratory manages facilities for thegeneral cultivation of plants and the rearing of several animal
species that do not qualify for housing in other facilities.
The Plant Culture Laboratory equips and manages approximately 75 culture boxes or growth chambers and 13
rooms with the P1P physical containment level required
for established and emerging model plants, such as thale
cress (Arabidopsis thaliana), rice (Oryza sativa), moss
(Physcomitrella patens), liverwort (Marchantia polymorpha), green alga (Chlamydomonas reinhardtii), and several
other flowering plants including carnivorous plants. The
facilities are also used to grow the sea anemone Exaiptasia
pallida. The culture space is used throughout the year by >70
researchers from external and internal groups.
In addition to regular culture conditions, extreme environmental conditions in terms of light and temperature areavailable for experiments. Three chambers (3.4 m 2 each)with controllable CO2, humidity, temperature, and light (max70,000 lux) conditions are also available. A tissue culture
rack with dimmable LEDs and pulse-width modulation con-99

National Institute for Basic Biology

Research Support

trollers is available for algae cultures that must be exposed toprecise light concentrations. Autotrophic and heterotrophic
culture devices are also available for researchers working
with cyanobacteria, algae, and cultured flowering plant cells.
Aseptic experiments can be performed in an aseptic roomwith clean benches and a safety cabinet. Various analytical
instruments, including three flow cytometry systems and a
DUAL-PAM system to measure DNA content and chlorophyll fluorescence, respectively, are also available. A liquidhandling system for fully automated and simultaneous insitu hybridization of sections of up to 60 glass slides is alsoprovided.
There is also a 386 m2 experimental farm next to the NIBB
Myodaiji area building for maintaining Japanese morning
glories and related Ipomoea species, several carnivorous
plants, castor beans, and other flowering plants that must be
cultivated outdoors. Three heated greenhouses (measuring44, 44, and 45 m2) contain sensitive carnivorous plants andthe periodically mass-flowering plant Strobilanthes flexicaulis. Four air-conditioned greenhouses (4, 6, 9, and 9 m2)
are provided for the cultivation of Japanese morning glories
and additional carnivorous plants. Two of the air-conditioned
greenhouses (9 and 18 m2) meet the P1P physical containment level and are available for experiments using transgenic rice plants, Japanese morning glories, and carnivorous
plants. The Plant Culture Laboratory also maintains a 46
m2 building with a storage area and workspace. Part of thisbuilding is used for rearing Japanese rhinoceros beetles andthe common grass yellow butterfly.
Between April 2021 and March 2022, a ploidy analyzer wasintroduced. It is a compact flow cytometer that is suitablefor ploidy and genome size analyses of plants, animals,and microorganisms. The liquid handling system has been
updated to support whole-mount in situ hybridization.
There are two groups in the Laboratory of BiologicalDiversity, and the assistant professors at this facility, Dr. A.
Hoshino and Dr. K. Tsugane, are the principal investigatorsof each group. For additional details, please refer to the laboratory’s webpage.

Figure 5. Newly introduced ploidy analyzer.

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CELL BIOLOGY RESEARCH FACILITY
Associate Professor:

WATANABE, Eiji

The Cell Biology Research Facility provides various equipment for tissue and cell culture. This laboratory is equippedwith safety rooms which satisfy the P2/P3 physical containment level, and is routinely used for DNA recombination
experiments.

Figure 6. Equipment for P3A experiments