Designing a Plant Tissue

Culture Laboratory

Dr Fatimah Kayat

Outline

Introduction

General laboratory design

Washing & Storage

Media Preparation

Instrumentation

Aseptic transfer area

Primary growth room

INTRODUCTION Certain elements are essential for a

successful operation

The correct and strategic design helps to achieve a high standard work.

Careful planning is an important step considering the size and location.

A small lab should be set up first until the proper techniques and markets are developed.

STRATEGIC PLANNING

Good locations

Isolation from traffic.

No contamination from adjacent rooms.

Access to water & electricity supply

Connection to a septic system or sanitary sewer

CONSIDER

CONSIDER

Water service

Air conditioning

maintain constant temperature

Electrical service capacity

equipment, lights and future expansion

An emergency generator

to operate essential equipment during power breakdown

PRIORITY

Cleanliness No contamination.

Easy to wash walls and floors.

High efficiency particulate air (HEPA) filters.

An enclosed entrance, sticky mats, no shoes.

Traffic pattern and work flow The cleanest rooms or areas

Aseptic transfer area

Primary growth room

Could not enter directly from outside.

Should be enclosed with doors leading to each other

Media preparation area, washing area, storage

area should be outside these rooms.

Media preparation area should lead to the

sterilization area

TO CONSIDER

Washing area located near sterilization and media

preparation area.

should be equipped with at least one large sink.

The pipes can be PVC to resist damage from acids

and alkaline

Hot/cold water sources

Water distillation and/or deionization devices

Ovens or hot air cabinets (75 oC) close to the washing

and storage area

WASHING & STORAGE AREA

Storage area should be close to the washing

area for storage

Need to be accessible to the media preparation

area

Dust-proof cabinets can be used in the storage

area

WASHING & STORAGE AREA

GlasswareGood quality of borosilicate glass

Highly resistant to autoclaving, breakage and scratch proof

E.g: Pyrex, Corning, Kimble, Schott

Water source and storage area should be

convenient to the media preparation area.

A variety of equipment for media preparation are

generally located in the media preparation area.

MEDIA PREPARATION &

STERILIZATION AREA

BASICS EQUIPMENT

Refrigerator/freezer to store chemicals and stock solutions

(usually in media preparation area)

High quality water Purchasing bottled water is not economical for mass production

Distilled water, double distilled water devices (usually in washing area)

A triple beam balance

Useful for large amounts over 10 grams

Can measure down to 2 mg is essential.

Balances

a microbalance

a less sensitive top loading balance

BASICS EQUIPMENT

Hot plate/stirrer

At least one hot plate with an automatic

stirrer is needed to make semi-solid media.

BASICS EQUIPMENT

pH meter

to measure media pH.

Some laboratories use pH indicator paper.

BASICS EQUIPMENT

Aspirator or vacuum pump

to filter sterilization of chemicals

also used to disinfest plant material

Other Equipment

Autoclave or Pressure Cooker

vital part of a laboratory

to sterile media, apparatus, used vessel

Self generating steam autoclaves are more

dependable and faster to operate.

BASICS EQUIPMENT

Optional equipment

Microwave oven are convenient for defrosting

frozen stocks and heating agar media

Dissecting microscopes are useful for

dissecting floral and shoot apices, and

observing plant culture growth

Labwashers/dishwashers are useful for

washing and drying glassware (mass

production)

OTHER EQUIPMENT

Laminar flow hood

For sterile working procedure

With High Efficiency Particulate Air (HEPA)

filter – filter the air to high degree

With UV germicidal lamp – sterile the shell and

content when not in use

Horizontal laminar flow unit---An enclosed work area that has sterile

air moving across it. The air moves with uniform velocity along

parallel flow lines. Room air is pulled into the unit and forced through

a HEPA (High Energy Particulate Air) filter, which removes particles

0.3 μm and larger.

BASICS EQUIPMENT

LAMINAR FLOW HOOD

Incubator

To incubate the culture

Controlled conditions (temperature, light,

humidity)

OTHER EQUIPMENT

Need to be as clean as possible

Preferably separated from other rooms

to decreases spore circulation and allows personnel to

leave shoes outside the room.

Laboratory shoes and lab coats should be worn

Aseptic working area. Ultraviolet (UV) lights are sometimes installed to

disinfect the room (only switched on when peopleand plant material are not in the room)

Safety switches can be installed to shut off the UVlights.

air filters are sometimes installed to provide clean air

ASEPTIC TRANSFER AREA

Smooth surfaces inside the aseptic transfer area to minimize the amount of dust and easy to wipe.

Several electric outlets are to be installed to accommodate Flow hoods,

Hot bead sterilizer

Microscopes.

ASEPTIC TRANSFER AREA

ASEPTIC TRANSFER AREA –

commercial production

PRIMARY GROWTH

ROOM

Should consider:

temperature

relative humidity

lighting units and shelves.

Varies depending on

the size of the growth room

its location

the type of plants grown

Temperature is the primary concern It affects decisions on lights, relative humidity, and

shelving.

Generally, temperatures are kept at 24 ±1 oC.

Cooling the room is usually a greater problem

than heating;

cooler T can be obtained by installing heat pumps,

air conditioners or exhaust fans.

Small fans can be placed at the ends of shelves to increase air flow and decrease heat accumulation.

PRIMARY GROWTH ROOM

PRIMARY GROWTH ROOM

Shelving within primary growth rooms can vary

depending upon the situation and the plants

grown.

Wood is inexpensive and easy-to-build shelves.

It could be particleboard or plywood and painted

white to reflect the room's light.

Expanded metal is more expensive but provides

better air circulation.

Some plant cultures can /need to be kept in

complete darkness

Most culture rooms are under light condition

Depending on

plant species

developmental stage

Automatic timers

help to maintain desired photoperiods

Reflectors can replaced bulbs

reduce cost

Heat generated by lights may cause

condensation and temperature problems.

PRIMARY GROWTH ROOM

Relative humidity (RH) is difficult to control

inside growing vessels

fluctuations in the culture room may have a

deleterious effect.

Cultures can dry out if the room's RH is less

than 50%;

humidifiers can be used to correct this problem.

If the RH becomes too high, a dehumidifier

is recommended.

PRIMARY GROWTH ROOM

PRIMARY GROWTH ROOM

PRIMARY GROWTH ROOM

PRIMARY GROWTH ROOM

Any questions?