Group 1 : Aseptic Work Area

Teh Ai Yeen Gan Soon Chong Sivakanth Selvamani Rubananthan Ramasamy Jegatheeswaran Krishnan 260110103004 260110103008 260110103015 260110103021 260110103026

Introduction : Tissue Culture

A method of biological research in which fragments of tissue from an animal or plant are transferred to an artificial environment in which they can continue to survive and function. Facilitated via use of a liquid, semi-solid, or solid growth medium, such as broth or agar

 Involves exposing tissue to a specific regimen of nutrients, hormones, and light under sterile, in vitro conditions to produce many new clone of the original mother, over a very short period of time Involve 3 stages : Initiation Multiplication Root Formation

Stage 1

It concerns the establishment of plant tissue in vitro by sterilising the material and initiating it into culture.

Stage 2

The in vitro plant material is re-divided and placed in a medium with plant growth regulators that induce the proliferation of multiple shoots. Repeated many times until the number of plants desired is reached.
Involves the introduction of hormones to induce rooting and the formation of complete plantlets.

Stage 3

Aseptic Work Area

Successful cell culture depends heavily on keeping the cells free from contamination by microorganisms such as bacterial, fungi, and viruses. Nonsterile supplies, media, and reagents, airborne particles laden with microorganisms, unclean incubators, and dirty work surfaces are all sources of biological contamination.

 Aseptic technique provide a barrier between the microorganism in the environment and the steril cell culture. Cell culture hood is normally used. Cell culture hood reduce contamination from aerosol and airborne particles.

Cell Culture Hood

 Air is drawn through a HEPA filter and blown in a very smooth, laminar flowtowards the user. The cabinet is usually made of stainless steel with no gaps or joints where spores might collect. Such hoods exist in both horizontal and vertical configurations, and there are many different types of cabinets with a variety of airflow patterns and acceptable uses.

 The cell culture hood should be properly set up and be located in an area that is restricted to cell culture that is free from drafts from doors, windows, and other equipment, and with no through traffic. The work surface should be uncluttered and contain only items required for a particular procedure; it should not be used as a storage area.

 Before and after use, the work surface should be disinfected thoroughly, and the surrounding areas and equipment should be cleaned routinely. For routine cleaning, wipe the work surface with 70% ethanol before and during work, especially after any spillage.

 You may use ultraviolet light to sterilize the air and exposed work surfaces in the cell culture hood between uses. Using a Bunsen burner for flaming is not necessary nor is it recommended in a cell culture hood. Leave the cell culture hood running at all times, turning it off only when they will not be used for extended periods of time.