I'm working on hairy root culture in Mucuna pruriens with two strains MTCC and KCTC which are very old cultures. I'm unable to induce hairy root culture.
Any one is working on same plant?
Thank you
Can you send me your protocol in which you were not able to get induce roots?
Also, more details about your A. rhizogenes strains might be helpful. There are a number of MTCC and KCTC stains, and not knowing the exact strain makes it more difficult to find relevant literature that might help fix your problem.
@S. Wenkart and @Bekele Abebie thank you for your response.
The protocol is,
The strain was inoculated for 48 hours, then OD was checked.
Wounding was done and broth was poured and left for infection to occur for 45 min.later excess broth was removed by filter paper and placed in MS media at dark for 2 days and then media was changed. Ms media with antibiotics was taken and cultured there.
@ Ray Collier im so sorry that I do not have much details of the strain as it was a very old one present in my lab which was not maintained
There are indeed many strains. I still have one left in my lab of the so called natural ones. Which I believe are stronger. One can order them in US. Hope I dont loose it. Ok of course I keep a replica in minus 70C . But I like better to transfer to fresh plates once a month.
Rakesh,
If I understand correctly, you have directly exposed the plant tissue to the growth medium for the Agrobacterium, yes? If this is the case, when you repeat the experiment, you might try pelleting the Agrobacterium cells by centrifugation, decanting the growth medium, then washing the cell pellet (I usually completely resuspend by rapid vortexing) with sterile ultrapure water (use a volume of sterile water equal to the volume of the culture), pellet the cells again, remove the wash water, then resuspend the cells in infection medium (lots of choices, this is an empirical part of your work) to the appropriate optical density (I like 600 nm, other people use different wavelengths) and you can proceed to infection. The reason I am suggesting this strategy is because you might be killing plant cells with the antibiotics, washing the cells can help mitigate this problem.
Ray Collier
Dear sir,
I Jaspreet, pursuing PHD in plant tissue culture and also studying agrobacterium mediated transformation with MTCC 2364 and MTCC 532 strain. I am also unable to induce hairy roots in my culture. Recently i just saw your answer on research gate..>>
Can u send me proper protocol which u have mentioned in your answer or any reference.
With regards
Jaspreet Kaur
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