Hi guys! I'll be blogging on something that we do not usually have the chance to do in our curriculum! During this period, I'm like a gardener, the only difference is that I'm working in a plant lab and not a garden.. :)
Okie! Now about plant subculturing...
A subculture is actually a culture made by transferring tissue from a previous cultures to a fresh medium in order to prolong the life of a particular strain where there is a tendency to degeneration in older cultures. A plant subculture in many ways are similar to a cell subculture in terms of aseptic techniques involved, characteristic of a subculture and precautions to be taken.
Before a subculture can be carried out, the media used should be autoclaved at 121 deg cel for 15 min in order to kill off any microorganisms. After the media is autoclaved, the container should not be open unneccessary as it is prone to contamination. If necessary, it can only be opened in a horizontal flowhood which confers more product protection.
The selected plant culture has to be in a healthy state and free of contamination. The whole subculturing process have to be done in the horizontal flow hood to minimize contamination. Forceps and scissors should all be sterilized before use. The flow hood should also be UV and swapped with 70 % ethanol before use. Strict aseptic techniques applies. The containers is opened by applying force onto the four edges of the containers with thumbs and index fingers only. Once the plant container is opened, hands with gloves should not be placed over the plant culture. Only sterilized scissors and forceps are allowed to touch any of the plant culture, media or within the containers. The lower portion of the stem are cut and retrieved out using a forcep. All the leaves are then removed, using 2 forceps. Hence by removing the leaves, the nod of the stem is exposed in which new plantlets will grow from them. The stem is then inserted lying horizontally into the media with the use of forceps. the stems are aranged in organized manner (in rows) so as to prevent overcrowding.
Finally the plant subculture is ready for growth in the growth room. Nevertheless, the plant subculture are supplied with 12 hours of light daily and should be constantly check for contamination. Once sufficient plant subculture is grown, it is havested for use.
feel free to ask any question!
Royston Tan
0503289A
8 comments:
Hey
How are Forceps and scissors sterilized? See ya =)
Eugene Wong
TG02
Hey
Just wondering...what are the differences between a 'horizontal flowhood' and the normal laminar flow cabinets that we use?
Thanks
Azhar
TG01
heya...wanna ask u....is there a maximum no. of times u can subculture the plants? and when u keep subculturing...will it affect certain characteristics of the plants???
zahirah tg02
Hi eugene,
The forceps and scissors are sterilized by a glass bead sterilizer which sterilized the instruments at 250 degree C and the glass beads is to maximise the conduction of heat to all parts of the instruments.
Hi azhar,
Laminar flow cabinets consist of both horizontal and vertical flow hood. While another class of cabinets are known as biosafety cabinets.
Overall, laminar flow cabinets, either vertical or horizontal, may be used to enhance sterile conditions required for cell culture procedures. The choice between the two types may be made on the basis of technical differences (in ensuring continuing high air quality), size and configuration of work area, and envisaged techniques to be used. They both offer no protection to the operator and environment.
For more info, you can check out http://www.adelaide.edu.au/mbs/safety/biosafety/cabinets.html#Hlamflow
Hi zahirah,
From what i have learnt subculturing of plants can lasts as long as two years with minimum effects on the plants characteristics. In reality, the plant subculture do not lasts that long as the plant culture are prone to contamination and are disposed appropiately once contamination is observed.
However, when hormones are used in the media, the plants will become more dependant on the hormones to stimulate growth.
Hope that helps!
Royston Tan
TG01
0503289A
hey...
so what do you do to those removed stems and leaves? throw them away? are these subcultured plants used anywhere, like TP's greenhouse?
Ying Ying
TG01
Hi ying ying,
The stems with exposed nods are used for subculturing while the removed leaves are disposed appropiately. Normally, these plant tissue culture are cultured in the growth room but they can also be cultured in TP greenhouse. They are cultured in growth room as the plant tissue culture are prone to contamination. Growth room is a clean room which will minimise the chances of contamination and it is also a controlled environment where growth of the plantlets can be optimised unlike the harsh environment of TP greenhouse.
Hope these helps =D
Royston Tan
TG01
0503289A
hey hi
you mentioned that " Once sufficient plant subculture is grown, it is havested for use.
" usually how long does it take for the plants to grow?? how long u all culture the them for? definite or indefinite period?
Vino
plant tissue keep in glass bead sterilizers to see the growth in research lab.
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