contam or sediment? - Advanced Mycology

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Agar, Petri Dish

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down44
Johnson

Registered: 12/01/04
Posts: 177
Last seen: 7 months, 23 days

contam or sediment?

#9896699 - 03/02/09 12:20 PM (3 years, 2 months ago)

my friend tried to clone a fruit by taking a stem tissues sample, he took 2 diff samples in 2 diff jars its been a week and he has noticed a sediment floating ramdomaly and pieces sticking to the tissue samples that has a rust colored tint to them. Should he discard and try again or give these lc's a try?

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fastfred
Old Hand

Registered: 05/17/04
Posts: 6,242
Loc: Dark side of the moon

Re: contam or sediment?

[Re: down44]
#9897755 - 03/02/09 03:22 PM (3 years, 2 months ago)

Discard them. It doesn't sound like you used proper procedure. Cloning straight to LC is very prone to failure if you don't do it right.

It's much better to put your tissue on agar and then transfer a clean piece to LC.

-FF

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down44
Johnson

Registered: 12/01/04
Posts: 177
Last seen: 7 months, 23 days

Re: contam or sediment?

[Re: fastfred]
#9900106 - 03/02/09 08:39 PM (3 years, 2 months ago)

thanks fred

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fastfred
Old Hand

Registered: 05/17/04
Posts: 6,242
Loc: Dark side of the moon

Re: contam or sediment?

[Re: down44]
#9903535 - 03/03/09 12:30 PM (3 years, 2 months ago)

To be more specific... I really have never gone straight to LC. I always grow things out in petri dishes first. The procedure I use results in about a 75%+ success rate as far as contaminants per tissue sample.

What I do is take a stem and cut a length out of it. Then I dip it in straight peroxide for about 5 min. Then I dip it in 10% bleach solution for about 5 min. Then I cut it up and place about 4 pieces per dish. Then I pick out the uncontaminated pieces and transfer to another dish.

That works plenty well for wild specimens. If I were trying to go straight to LC I would do the same procedure but I would cut away the surface bits and get some inner tissue to start the LC. You might even subject the inner tissue to the same bleach/peroxide treatment. The important aspects are to properly sterilize and flame your scalpel after each cut.

-FF

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