The plant which I am planning to do in-vitro propagation is Sarcostemma. Can I use its internodal and nodal part as explant material? Does its latex cause any contamination or browning of medium?
Browning of medium is of no concern until it hampers callus induction, just ensure surface sterilization, use 70% alcohol followed by Hgcl2 apart from fungicide ttreatment. In case of contamination (Endophytic), u can go with streptomycin added medium.
Internodal and nodal segments of laticiferous plants can be cultured in vitro. The latex found in the explant will not hamper the explant growth under in vitro. Proper surface sterilization after removal of dead scales of axillary buds with fungicide followed by mercuric chloride has to undertaken to prevent contamination of the explants. Take the axillary buds from the shoots not more than 3 weeks which shows no lenticels development on its bark which is the hiding site for microbes.
Apart from taking above mentioned precautions, if still browning around ex plant occurs, the ex plants can be shifted to fresh place within the same bottle. By doing this process few times the browning can be tackled and eventually reduced.
Over and above the already mentioned precautions , you can try addition of activated charcoal in the medium, provided charcoal doesn't otherwise affect the development of your plant material. In our experience, it has been observed that charcoal does help very much in preventing toxic exudates affecting the cultures.
Yes, proper surface sterilization steps as mentioned by others are important to avoid contamination. Use of activated charcoal (AC) at an optimum concentration is really an effective way of controlling exudates and browning of explants. Please optimize the concentration of AC concentration for optimum result.
Besides what has been mentioned by others on this thread, those who suggest Activated Charcoal(AC) must specify which charcoal they are referring to? acid,basic, phosphate free etc. Any attempt to see any Chemical Catalogues would refer to many types. My own experience is neutral grade( about 30 yrs ago I relied on Darco G 60) very effective for initiating cultures and rooting studies in vitro for variety of Tree tissue culture studies.
Yes, charcoal can help. It has to be the washed one, acidfree. And I use only 0.02% in order to have some effect of the charcoal, but that would not absorb hormones etc from the medium.
You can also try with adding ascorbic acid in the medium. also keep removing the explant at different places in the same medium
As mentioned, antioxidants like ascorbic acid or picric acid, adsorbants like activated charcoal, and frequent subculture should be able to control browning of the explants. Use activated charcoal with caution since it will also adsorb the phytohormones in the media, so the effective (available) concentration will be less than what you put in. Also, cut off the browned portion of the explant before subculturing. The amount of exudates and browning should also decrease with successive subcultures. A combination of these should help you preventing the browning of the explants.
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