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InvisibleDoc_T
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Micropropagation at home.
    #14013111 - 02/23/11 11:53 AM (12 years, 11 months ago)

Ok, I was reading here about micropropagation the other day, and wondered why it's so fancy and exotic. Why we don't have home teks.
A few PMs later, I'm convinced it can be done.

Well, so today I go down to the local garden shop, I'm looking at the shelves and this employee asks if I need help.
"Rooting hormone?"
"What are you rooting," she wants to know.
"I was reading on the 'net about growing plants in dishes, rather than from seed. Sounded so cool I had to try it."
"Oh, you want this." turns around, grabs a bottle off the shelf, hands it to me.

"So this works?"
"Oh, yeah, I did it college," she says "it was one of my favorite experiments."
"Really? So, I need plant food too right"
"Yep. Miracle Grow." Hands me the MG.

"Ok, so... do I want silica gel? Or agar?"
"Agar." No hesitation, no uncertainty.
"Agar."
"Agar."

"So, this really works? For reals?"
"Oh yeah, works great"

So I guess I'm going to try it. :shrug:
Plan for the first run is to mix the MG and root hormone at whatever concentration the labels says is normal, use that to make agar jars.
I'll be using PF jars with plastic lids.

Ideally, one would like a plant with a known sex, but I just have some seeds.
It's 50/50 odds, right? If I sprout three seeds, I should get at least one female?


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Edited by Doc_T (02/23/11 12:05 PM)


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OfflineEl Douche
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Re: Micropropagation at home. [Re: Doc_T]
    #14013232 - 02/23/11 12:22 PM (12 years, 11 months ago)

I use rooting hormone for house plants. I just cut off a small branch, pluck the bottom leaves, dip the stem in hormone powder - including at least one nub (branching point), and place in tap water. Roots usually grow out in a week or 2. You could also place the dipped cutting in potting soil or other media and put a humidity tent over it untill the plant grows roots (you can use an inverted cut 2-liter as a tent).

I don't know how this will work with your specific plant, as my described method does not work on all of my plants.

You could put some Miracal Grow in the water.

I would think that agar would be difficult because it is prone to contaminants. The hormone used to come with fungicide, but the last time I looked it could not find the combination.

The hormone is toxic, so don't breath the dust or get it on your skin. A very small amount will kill an animal, and it is used in small amounts to kill fish in lakes.


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Trade List : Wanted/have edible cultures, ethobotanicals, cool plants, cacti.


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Re: Micro propagation at home. [Re: El Douche]
    #14013387 - 02/23/11 12:54 PM (12 years, 11 months ago)

sterilisation is very important once you've got that sorted then you should be ok. Ive been waiting for this, will see if it works compared to the normal route pretty much everyone Ive read up on has used which is getting all the chemicals and making their own media up and using M&S media


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Re: Micro propagation at home. [Re: Djcorbetto]
    #14013413 - 02/23/11 12:59 PM (12 years, 11 months ago)

Sterile technique I have down, it's fundamental to mycology. :thumbup:

Completely unrelated question, there was a cactus name Trichocereus transchekii, or similar. Is that one I want to back and buy?


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Re: Micro propagation at home. [Re: Doc_T]
    #14013460 - 02/23/11 01:08 PM (12 years, 11 months ago)

Done a bit of googling at Trichocereus terscheckii seems to be the one. on erowid it says that anyway.


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Re: Micro propagation at home. [Re: Djcorbetto]
    #14013478 - 02/23/11 01:11 PM (12 years, 11 months ago)

That's a good one?!?!

Edit- apparently nonmagical but a fun cactus anyway.
I may get it if it's there next week, but not worth driving back for.


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Edited by Doc_T (02/23/11 01:20 PM)


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Re: Micro propagation at home. [Re: Doc_T]
    #14014039 - 02/23/11 02:33 PM (12 years, 11 months ago)

If it doesnt work, try using the hormones at way lower conc than suggested on the label.

You'll need to look at the active ingredient (probably IAA, NAA, IBA or a combination) and then do some calcs to bring the level to µM levels...

try it out. you have sterile technique down, you shouldnt have a problem.


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Re: Micro propagation at home. [Re: caricapapaya]
    #14014054 - 02/23/11 02:36 PM (12 years, 11 months ago)

Label says IBA. :thumbup:
I used just a knifetip of it, my scale won't measure this small.
Anybody know whether the stuff is even PCable?


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Re: Micro propagation at home. [Re: Doc_T]
    #14014333 - 02/23/11 03:08 PM (12 years, 11 months ago)

yes, it can be autoclaved.

also, if you can dissolve it something, you can make a concentrated stock by weighing out more than you need, and then dilute that by volume instead of weight


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Re: Micro propagation at home. [Re: Doc_T]
    #14014348 - 02/23/11 03:10 PM (12 years, 11 months ago)

research into forming callous cultures, these can be split over and over to make hundreds of cultures, they grow under a specific hormone(i can't remember which now)

then you chance the most proeminent hormone on the cultures and they will form plantlets, which can be extracted from the agar just as normal seedlings.

i had some papers on it, i'll see if i can find them.

also, look into murashige skoog medium, it's a very basic 'good for all' medium.


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Re: Micro propagation at home. [Re: brshroomer]
    #14014371 - 02/23/11 03:14 PM (12 years, 11 months ago)

Quote:

brshroomer said: murashige skoog medium, it's a very basic 'good for all' medium.




I'm hoping this will be a local-ingredients grow. I don't want to send out for anything.
Several reasons, biggest being money. I'm just goofing around here, I don't plan to really grow anything much.
So sending out for supplies makes no sense.

But also I just think it's cooler if you can do it with regular OTC stuff.


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Re: Micro propagation at home. [Re: Doc_T]
    #14014430 - 02/23/11 03:21 PM (12 years, 11 months ago)

i agree that OTC ready mixed nutrients are WAY cheaper and practical, but i have to test them yet, i don't know if the proportions in the solution are right.

if it does work that may be a breakthrough for home tissue culture.

also, consider adding dextrose to the medium, it is added in MS medium, and it's gotta be there for some reason.


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Re: Micro propagation at home. [Re: brshroomer]
    #14014455 - 02/23/11 03:24 PM (12 years, 11 months ago)

Oh, I wondered about dextrose. :frown: Next time maybe.
I'm sprouting two seeds, that gives me 75% chance of at least one female.


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Re: Micro propagation at home. [Re: Doc_T]
    #14014543 - 02/23/11 03:36 PM (12 years, 11 months ago)

Sucrose is better from what I have read...


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Re: Micro propagation at home. [Re: Burbles]
    #14014548 - 02/23/11 03:37 PM (12 years, 11 months ago)

Table sugar? Really? Weird.


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Re: Micro propagation at home. [Re: Doc_T]
    #14014665 - 02/23/11 03:57 PM (12 years, 11 months ago)

Yes - this actually makes sense, that is the main sugar used within plants, and that also explains why sucrose is so available, we get it from plants.
Other than that, I don't know why, but it is what I have read.

I was thinking today in class that propel - that gaterade stuff, might actually be a good thing to add to these experiments, as it has EDTA - which is a common chelating agent and would help the metals stay in solution better, in addition it also has small amounts of various B vitamins (some are needed by plants, IIRC - like thiamine?) and has other things like citric acid (its used in MicroP for various reasons, as a nutrient, pH regulation, etc) and ascorbic acid (another chelating agent, if I remember correctly - though probably would decompose on PCing)

Might be worth a shot, I don't know.. I think EDTA is important though, and is hard to find OTC unless it is in one of those gunky EDTA supplements...

Also:
Link & description about Banana micropropagation and other exotic fruit/plants
http://www.thelaboratory.org/talk/index.php?topic=134.0
Amamature Micropropagation thread in general.. has some useful info...
http://www.thelaboratory.org/talk/index.php?topic=18.0
First attempt at micropropagation... using potato cuttings, guano tea
coconut and sea water
http://www.thelaboratory.org/talk/index.php?topic=202.0
And also there is a bit more about growing out cacti callus cultures, etc.. but i don't want to spam my site - but there is a pretty good amount of micro/plant tissue culture info there
Also check out these micropropagation videos, great for beginners using the Home plant tissue culture materials..
http://www.youtube.com/user/fbt2007

glad you are trying some stuff out!


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Re: Micro propagation at home. [Re: Burbles]
    #14014710 - 02/23/11 04:06 PM (12 years, 11 months ago)

good luck! :thumbup:

IBA stands for indole butyric acid, not bad rooting hormone at all :thumbup: if you've got powder form it's better 'cos solutions don't store for very long even if refrigerated


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Re: Micro propagation at home. [Re: cc2]
    #14014768 - 02/23/11 04:16 PM (12 years, 11 months ago)

Quote:

GIBBERELLIC ACID (GA3)

Stock solution of gibberellic acid: I ,000 ppm
1. Weigh 0. 2 p of gibberellic acid and dissolve well with some alcohol drops.
Add 200 ml of distilled water
2. Keep in a conveniently labeled vial at 0°C.
The gibberellic acid may be sterilized together with the culture medium: however, the loss of some activity is also possible.
One ml of concentrate solution (1,000 ppm) contains 1 my of gibberellic acid.

NAPHTHALENE ACID (NAA)

1 Stock solution of NAA 1,000 ppm
Weigh 0.2 g of NAA and dissolve well with some NaOH 1N drops.
2. Add 200 ml of distilled water.
Keep it in a conveniently labeled vial at 0~O.
One ml of stock solution (1,000 ppm) contains 1 mg of NAA.

N-6-BENZYLAMINOPURINE(BAP)

Stock solution of BAP: 1,000 ppm
1. Weigh 0. 2 g BAP and dissolve well with some drops of NaOH 1N.
Add 200 ml distilled water.
2. Keep in a conveniently labeled vial at 0°C.
BAP may be sterilized together with the culture medium; however, the loss of some activity is also possible.
One ml of stock solution (1,000 ppm) contains 1 mg of BAP

INDOLEACETIC ACID (IAA)

Stock solution of IAA: 1,000 ppm
1. Weigh 0.2 mg of IAA and dissolve well with some alcohol drops.
Add 200 ml of distilled water.
2. Keep it in a conveniently labeled vial at 0°C.
Sterilization by filtration is recommended.
One ml stock solution (1,000 ppm) contains 1 mg of IAA.

KINETINE (KIN)

Stock solution of KIN: 1,000 ppm
1. Weigh 0.2 g KIN and dissolve well with some drops of NaOH 1N. Add 200 ml of distilled water.
2. Keep in a conveniently labeled vial at 0°C.
KIN may be sterilized together with the culture medium; however, the loss of its activity is also
possible.
One ml of the stock solution (1,000 ppm) contains 1 mg of KIN.

2,4-DICHLOROPHENOXYACETIC ACID (2,4-D

Stock solution of 2,4-D: 1,000 ppm
1. Weigh 0.2 g of 2,4-D and dissolve well with some alcohol drops. Add 200 ml of distilled water.
2. Keep in a vial conveniently labeled at 0°C.
2,4-0
may be sterilized together with the culture medium; however, a loss of its activity is also
possible.
One ml of the stock solution (1 000 ppm) contains 1 mg of 2,4-0.





The actual water solubility of many of these compounds is very low - and sometimes will not fully dissolve in water if you don't use the above methods.


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Re: Micro propagation at home. [Re: Burbles]
    #14015155 - 02/23/11 05:29 PM (12 years, 11 months ago)

I had a recipie for shoots for cannabis, and it just said: M&S media with 15g/litre of sucrose. when i spoke to carol at home tissue culture she reccomended that i got the M&S media with vitamins as its same price but better, ive read that if you add a multivitamin tablet to the solution that helps too


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Re: Micro propagation at home. [Re: Djcorbetto]
    #14015699 - 02/23/11 06:57 PM (12 years, 11 months ago)

It didn't have any hormones at all??
And it was for shoot formation, and not root formation?

The root formation would make a ton more sense to me, they root naturally - but it wouldn't make sense to have it grow more shoots and no roots if it is just a small random piece of flesh...

But I guess if it were a stem, it could easily grow both, esp. if it had a tip that would allow for leaves to grow out already..


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Re: Micro propagation at home. [Re: Burbles]
    #14017809 - 02/24/11 01:20 AM (12 years, 11 months ago)

I may have written it down wrong, infact im sure i remember seeing different hormones because it provoked me to look all over google and ebay to see how much they would cost me to purchase. I will look again and see if i can find the link for the page


EDIT: Found it

The media is M&S, with 15 g/L of sucrose, pH 5.8 before sterilizing (balanced with KOH) and 2mg/L kinetin, 0,2 mg/L IAA and 10 g/L agar.

it doesnt specify whether its for roots or shoots but then im still looking at his follow up as he got shoots on the plant material but so fare dont see a bit on roots.

Link: The media is M&S, with 15 g/L of sucrose, pH 5.8 before sterilizing (balanced with KOH) and 2mg/L kinetin, 0,2 mg/L IAA and 10 g/L agar.


i saw on that page there was a mention of a salvia recipe but they only put down what they had researched: "...Then placed in sterile glass culture tubes containing Murashige and Skoog medium supplemented with 2% glucose, Gamborg’s Vitamin solution, 0.2% Phytagel, and 5 μM 6-benzylaminopurine



Edited by Djcorbetto (02/24/11 01:30 AM)


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Re: Micro propagation at home. [Re: Djcorbetto]
    #14017892 - 02/24/11 01:48 AM (12 years, 11 months ago)

Oh and theres this:http://www.lycaeum.org/mv/anagrams/cannabis_tissue_culture.html


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Re: Micro propagation at home. [Re: Djcorbetto]
    #14020323 - 02/24/11 02:33 PM (12 years, 11 months ago)

Awesome, thanks for that link!


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Re: Micro propagation at home. [Re: Burbles]
    #14020503 - 02/24/11 03:06 PM (12 years, 11 months ago)

No problem, hopefully it helps you so you will make up a method of doing this which in turn will help me :smile:


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Re: Micro propagation at home. [Re: Djcorbetto]
    #14020951 - 02/24/11 04:44 PM (12 years, 11 months ago)



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Re: Micropropagation at home. [Re: Doc_T]
    #14034742 - 02/27/11 08:03 AM (12 years, 10 months ago)

Quick question about process:
I didn't have a plant handy so I'm sprouting some seeds.
I guess I need chlorophyll for this to work, not just a root tip?
But even a cotyledon should work, no need for a 'real leaf'?

Also, I don't think I can punch discs out of a leaf with a needle, I'm just going to cut tiny flakes with scissors.
I hope that's not cheating.


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Re: Micro propagation at home. [Re: Doc_T]
    #14034828 - 02/27/11 08:51 AM (12 years, 10 months ago)



--------------------
:atom: :tripmolecule: :lsdabc: :tripmolecule: :atom:


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Re: Micro propagation at home. [Re: doses]
    #14034851 - 02/27/11 08:58 AM (12 years, 10 months ago)

Quote:

doses said:
http://www.mediafire.com/?uy2jiizx2hz




Quote:

Moving the plantlets to regular soil
The key to sucessfully transferring your in-vitro plantlets into soil is to be very fastidious about washing off all the TC media from the roots.




Words of wisdom right there. Thanks!!!


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Re: Micro propagation at home. [Re: Doc_T]
    #14035704 - 02/27/11 12:24 PM (12 years, 10 months ago)

to cut the leaf up you could use a scalpel instead scissors


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Re: Micro propagation at home. [Re: Djcorbetto]
    #14036320 - 02/27/11 02:29 PM (12 years, 10 months ago)

Plants from a test tube... Its one of the best books out there...

Edit: Damn it, shroomery won't let me upload djvu files...

Here... http://www.thelaboratory.org/talk/index.php?topic=18.msg1285#msg1285


Edited by Burbles (02/27/11 02:31 PM)


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Re: Micro propagation at home. [Re: Burbles]
    #14036340 - 02/27/11 02:33 PM (12 years, 10 months ago)

Also you shouldn't need chlorophil at all.. if your medium has sucrose...the tissue callus does not do photosynthesis to any significant extent and could live in the dark if need be...AFAIK


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Re: Micro propagation at home. [Re: Doc_T]
    #14036356 - 02/27/11 02:35 PM (12 years, 10 months ago)

Quote:

Doc_T said:
That's a good one?!?!

Edit- apparently nonmagical but a fun cactus anyway.
I may get it if it's there next week, but not worth driving back for.






Supposedly it's active, but it's not mesc.


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Re: Micro propagation at home. [Re: HarveyWalbanger]
    #14036494 - 02/27/11 03:06 PM (12 years, 10 months ago)

hmm, lets see where this goes


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Re: Micro propagation at home. [Re: jokefox]
    #14036632 - 02/27/11 03:43 PM (12 years, 10 months ago)

I have no idea what's going on here...
So you are essentially trying to grow a plant, but only feeding off of the agar like a fungus? With the assistance of the plant food of course. I am curious to see your experiment summary.
GL


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Re: Micro propagation at home. [Re: Microppose]
    #14036655 - 02/27/11 03:47 PM (12 years, 10 months ago)

Quote:

Azurascender said:
I have no idea what's going on here...
So you are essentially trying to grow a plant, but only feeding off of the agar like a fungus? With the assistance of the plant food of course. I am curious to see your experiment summary.
GL





go type in micro propagation to google,
and you will be amazed


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Re: Micro propagation at home. [Re: jokefox]
    #14036785 - 02/27/11 04:10 PM (12 years, 10 months ago)

OMG, rather interesting. What kind of plant has he chosen? From what I understand, you get plant material and cell multiplication, but not the features of the plant?


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Re: Micro propagation at home. [Re: Microppose]
    #14036829 - 02/27/11 04:19 PM (12 years, 10 months ago)

basically you grow a plant from a tiny piece called a platlette or somthing ,
that calluses, the shoots & roots , then you have a clone plant of the original

theory is a thousand plants from a leaf


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Re: Micro propagation at home. [Re: jokefox]
    #14036897 - 02/27/11 04:30 PM (12 years, 10 months ago)



Or you could just grow the cells for alkaloid production, perhaps...
I.e grow root cells of sassafrass in a large liquid culture - its a bit different than true micropropagation, but the skills are the same.


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Re: Micro propagation at home. [Re: Burbles]
    #14037298 - 02/27/11 05:49 PM (12 years, 10 months ago)

Quote:

Burbles said:
Also you shouldn't need chlorophil at all.. if your medium has sucrose...the tissue callus does not do photosynthesis to any significant extent and could live in the dark if need be...AFAIK




this is ...awesome.

this could be a solution to mailing plants.


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Re: Micro propagation at home. [Re: Microppose]
    #14038105 - 02/27/11 07:57 PM (12 years, 10 months ago)

Quote:

Azurascender said:
OMG, rather interesting. What kind of plant has he chosen?




The kind I had a few seeds of laying around close at hand. :wink:

Yeah, theory is agar with plant food and 'stuff', grow clone plants from tiny shreds.

We'll see what we see.


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Re: Micro propagation at home. [Re: Doc_T]
    #14039345 - 02/27/11 10:32 PM (12 years, 10 months ago)

Ya'll have seen mushmush/Una's website right?

If not, your missing out.

http://www.mushmush.nl/?page=begin\gallery\tissue_culture

click through that inspirational stuff sometime..

I have been trying to figure out his "secret cactus culture' stuff and I am pretty sure I almost got it down - trick is coconut milk and a certain hormone or two.. .can't remember but I posted all the articles I found on cacti micropropagation on my site.

But this is the sort of stuff you see on his site:




I am mostly interested in the callus tissue culture -- imagine giving it just the right conditions to produce mescaline, grow faster, and just be an unidentifiable green blob :laugh:


Edited by Burbles (02/27/11 10:36 PM)


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Re: Micro propagation at home. [Re: Burbles]
    #14039990 - 02/28/11 12:32 AM (12 years, 10 months ago)

not just that, if it proves true that the plants need no light during micropropagation then this is some seriously important stuff.

you could send clones MUCH easier in the mail, i can think of a lot of plants that i would like to have but can't because they wouldn't withstand the trip(salvia for one)

even for cannabis as the OP is working with there is some utility, much better to send a green tiny blob in the mail than seeds. much lesser risk of crushing too.


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Re: Micro propagation at home. [Re: Burbles]
    #14040075 - 02/28/11 12:54 AM (12 years, 10 months ago)

Thread:



Small world.  My copy of Plants from test tubes arrived about a week ago. I have a month or so fantasizing about the uses of a kitchen tek for cannabis. So who wants to help write one? I found this research done on the best adjustment of the ms medium for hemp but am having trouble understanding exactly how to carry out the mixing without a firm background in chem.. Perhaps we can help each other.:rasta: Here is the linkhttp://www.pakbs.org/pjbot/PDFs/41%282%29/PJB41%282%29603.pdf
I want to revive some strains. Imagine the possibilities.

Op... Keep us updated.


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Re: Micro propagation at home. [Re: brshroomer]
    #14040100 - 02/28/11 01:01 AM (12 years, 10 months ago)

Quote:

brshroomer said:
not just that, if it proves true that the plants need no light during micropropagation then this is some seriously important stuff.

you could send clones MUCH easier in the mail, i can think of a lot of plants that i would like to have but can't because they wouldn't withstand the trip(salvia for one)

even for cannabis as the OP is working with there is some utility, much better to send a green tiny blob in the mail than seeds. much lesser risk of crushing too.





I think light is needed and depending on the developmental stage of the explant, the light could have disastrous effects. for instance if one were so inclined to send clones via mail in a ms medium, I assume the light cycle would need to be achieved with a 12 volt battery and a led; However I have no experience on how light effects plants in vitro...


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Re: Micro propagation at home. [Re: cacharstar]
    #14040530 - 02/28/11 07:01 AM (12 years, 10 months ago)

I got a PM asking for updates, but not much to say yet.
I checked my seeds last night. One had a root about an inch long, but hadn't gotten out of the seed hull.
When I picked it off, the seedling was discolored and looked icky. :frown:
Root seemed healthy, I put a few bits into a dish.
And a few bits of the leaf end went into another dish, just to see.

So I guess I still need a plant, I'm stuck at Step 0.


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Re: Micro propagation at home. [Re: Doc_T]
    #14040651 - 02/28/11 08:04 AM (12 years, 10 months ago)

I cant remember what plant you are trying to propogate, was it cannabis im a bit dopey at the moment and got a rubbish internet signal so cant go back. if it is cannabis do you not have any friends who live near you who grow or know someone who grows, and get a cutting from them. if you explain to them what you are doing then they might be more inclined to help you out and moeny always helps out. keep some of the plant to grow for a cutting so you have a reasonably steady source of plant material.


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Re: Micro propagation at home. [Re: Djcorbetto]
    #14040658 - 02/28/11 08:07 AM (12 years, 10 months ago)

Cannabis, yes. Yeah, I might be able to get a cutting from a friend of a friend. I can trade for it. :smile:

Other than salvia, what plants would home hobbyists want to grow this way?
Orchids, I know. What else? Cactus, maybe?


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Re: Micro propagation at home. [Re: Doc_T]
    #14040947 - 02/28/11 09:56 AM (12 years, 10 months ago)

Doc_T

did you sterilize the seeds?


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Re: Micro propagation at home. [Re: caricapapaya]
    #14040960 - 02/28/11 09:59 AM (12 years, 10 months ago)

Dipped in dilute bleach, like I usually do when germinating seeds. Then between wet paper towels in the dark.

I figure the leaves can be washed with sterile water, h202, whatever before rendering them for donor plugs.


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Re: Micro propagation at home. [Re: Doc_T]
    #14041044 - 02/28/11 10:26 AM (12 years, 10 months ago)

I thought you were germinating in vitro.

sterilizing tender new leaves can be problematic. not saying it cant be done, but its easier to sterilize seeds.


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Re: Micro propagation at home. [Re: caricapapaya]
    #14041149 - 02/28/11 10:48 AM (12 years, 10 months ago)

I could germinate on agar, I've got extra dishes on hand.

I think a new shoot is going to be basically sterile anyway, I don't see why there would be any contamination?


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Re: Micro propagation at home. [Re: Doc_T]
    #14041159 - 02/28/11 10:49 AM (12 years, 10 months ago)

.


Edited by HarveyWalbanger (03/07/11 02:50 PM)


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Re: Micro propagation at home. [Re: HarveyWalbanger]
    #14043596 - 02/28/11 05:16 PM (12 years, 10 months ago)




http://www.lycaeum.org/mv/anagrams/cannabis_tissue_culture.html


Cannabis tissue culture :laugh:


Why would the plant need light to grow if it is using the sucrose in the mixture? the callus tissue culture doesn't do a significant amount of photosynthesis, and light is primarily only used to aid in proper formation of the small plantelts... I.e grow up and not etioliate..

No need for light during the callus tissue formation AFAIK... and that would be the way you would want to send strains through the mail anyways for obvious reasons (decreased to no THC levels, unidentifiable, probably more hardy)


Edited by Burbles (02/28/11 05:22 PM)


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Re: Micro propagation at home. [Re: Burbles]
    #14043745 - 02/28/11 05:34 PM (12 years, 10 months ago)

Excellent post Burbles.
So this specimen in figure four, is after six weeks of growth on MS in a slant?


Edited by Microppose (02/28/11 05:35 PM)


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Re: Micro propagation at home. [Re: Burbles]
    #14043938 - 02/28/11 05:55 PM (12 years, 10 months ago)

Not sure how old the 4th picture is, the third  is 6 weeks, and I think the first two are only 3 weeks. The 4th picture is probably like 7-8 weeks, which seems kind of like a long time, but given this can be an exact clone of the plant you like, and you can do hundreds or thousands of them.. that isn't bad at all.

Also you can skip step 1-3. You really only need a small sterile shoot, which than it can be rooted.. this doesn't allow you to produce as many for obvious reasons - but as long as the cutting has a small place to grow new leaves/shoots - you could essentially use any piece of the plant.
That would easily result in hundreds from a good plant.
I haven't done much with micro-prop. Its my summer project though!

I will also be getting a ton of supplies, like ideal tissue culture jars (about 4 oz polypropylene, with a cap that functions like a petri dish cap... allowing for air flow but prevents contams.


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Re: Micro propagation at home. [Re: Burbles]
    #14052564 - 03/02/11 01:45 AM (12 years, 10 months ago)

well you need more then rooting and nutrients. unless you are just rooting cuttings essentially.  need to get a calus going of sort of stem cells take that get it to root and then to grow stem all using dif hormones which i dont think are easy to get ahold of as i tried a long time ago.  if there is a way let me know would be cool for orchids and stuff. just getting those dang hormones.  keep us updated if you find out if you can find them. be awesome send a callus culture of MJ much easier then a cutting    send a og kush wedge :smile:


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Re: Micro propagation at home. [Re: the man]
    #14052972 - 03/02/11 06:26 AM (12 years, 10 months ago)

I bought 25 grams of NAA and 5 grams of 6-BAP for about 10 bucks each I believe, many others are for sale as well... all over the place..


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Re: Micropropagation at home. [Re: Doc_T]
    #14055482 - 03/02/11 04:05 PM (12 years, 10 months ago)

Well fuck me running- this might work!

Hard to see in the pic, but I think I have some shoots growing out of root fragments.
Largest is 3-4mm, better pics should happen in a couple days.



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Re: Micropropagation at home. [Re: Doc_T]
    #14055595 - 03/02/11 04:30 PM (12 years, 10 months ago)

Shweet mang, what was your final medium mixture?


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Re: Micropropagation at home. [Re: Microppose]
    #14055949 - 03/02/11 05:31 PM (12 years, 10 months ago)

I would be shocked if growth were occurring that fast!


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Re: Micropropagation at home. [Re: Burbles]
    #14056061 - 03/02/11 05:52 PM (12 years, 10 months ago)

Quote:

Azurascender said:
Shweet mang, what was your final medium mixture?




MG/IBA agar, Miracle Gro and agar per label recipes with a pinch of IBA rooting hormone.
But that's not final, it's my first try.


Quote:

Burbles said:
I would be shocked if growth were occurring that fast!




It was a root segment, not a leaf.
But I'm still shocked- this idea might actually work.
Too early to call it a success, but it hasn't failed yet.

And contams don't seem to be a major concern, I had these in an incubator at 78F to encourage any bacteria to declare themselves.


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Re: Micropropagation at home. [Re: Doc_T]
    #14058126 - 03/03/11 01:40 AM (12 years, 10 months ago)

Have you taken any pictures of your attempt?


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Re: Micropropagation at home. [Re: Doc_T]
    #14058384 - 03/03/11 04:11 AM (12 years, 10 months ago)

Quote:

Djcorbetto said:
Have you taken any pictures of your attempt?




Only the one I posted 12 hours ago.

Quote:

Doc_T said:




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Re: Micropropagation at home. [Re: Doc_T]
    #14058418 - 03/03/11 04:43 AM (12 years, 10 months ago)

:popcorn:


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Re: Micropropagation at home. [Re: lucas_southoz]
    #14058422 - 03/03/11 04:47 AM (12 years, 10 months ago)

Somehow i managed to miss that pic, lets hope in a few weeks there will be some sign of life


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Re: Micropropagation at home. [Re: Djcorbetto]
    #14058433 - 03/03/11 04:57 AM (12 years, 10 months ago)

There's signs of life right now. Are you reading the thread?
Sorry it doesn't show in the pic, but there's a shoot growing in that jar.


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Re: Micro propagation at home. [Re: Doc_T]
    #14058644 - 03/03/11 07:38 AM (12 years, 10 months ago)

Im vaguely trying to read the thread just a bit busy in uni the past couple of days so not really had a chance to read it all. so did you grow the shoot from a seed placed in the jar? or was it sliced up?


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Re: Micro propagation at home. [Re: Djcorbetto]
    #14060971 - 03/03/11 03:59 PM (12 years, 10 months ago)

dude, it's all alredy in the thread, how about doing some reading?

doc, i'm glad to know it's working, that might be a important step for hobbyists.


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Re: Micro propagation at home. [Re: brshroomer]
    #14061459 - 03/03/11 05:09 PM (12 years, 10 months ago)

Hay guiz!  Look at the other jar!
This is officially a thing now. Next batch, I'm adding a bit of sucrose.
Green in the background is from the original leaf, but it never looked that good. Stem in front is growing straight up out of the agar.

Anybody got a spare cactus? Or seeds? I'd love to try this with a cactus. Switching to soil would be easy, it's super clean.



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Re: Micro propagation at home. [Re: Doc_T]
    #14064059 - 03/04/11 12:05 AM (12 years, 10 months ago)

Great work Doc. Have been poking my head in this thread and I now think it's time to congratulate you on your efforts.

I will continue to watch your progress.:sun:


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Re: Micro propagation at home. [Re: karode13]
    #14064711 - 03/04/11 04:46 AM (12 years, 10 months ago)

Thanks!
Like I said earlier, it's not 'success' yet. But it hasn't failed yet, so not bad for a first try.

Anybody else interested enough to play along at this early stage?
We should start comparing notes and recipes right away, narrow it down quickly.
One variable that might be relevant: gas exchange in the jars. Do we need filtered lids?


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Re: Micro propagation at home. [Re: Doc_T]
    #14064915 - 03/04/11 07:08 AM (12 years, 10 months ago)

Doc-

That looks good with no contams, the leaf pieces are still green (a good sign) as for the root segment....I dont see it looking like a shoot.

Not trying to get you down. There are a few ways that you can go from root segment to shoot :

Adventitious shoot formation or embryogenesis (new embryos from somatic tissue) are 2 of them. if a shoot starts to grow, it will typically be green (after a little while anyway, especially if exposed to light, and especially with no sugar in the medium).

as for fae, some plants are very sensitive, and need a vented conatiner, adn some dont care. most will grow better with vents (bigger leaves, thicker stems) but dont necessarily need it.

keep an eye on those peices, look for smooth globular structures (often the beginning of new embryos) or anything that looks leafy.


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Re: Micro propagation at home. [Re: caricapapaya]
    #14065025 - 03/04/11 08:04 AM (12 years, 10 months ago)

You only added IBA, correct?
Don't you also want something such as 6-BAP for it to grow into a callus or to aid in rooting/shooting?


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Re: Micropropagation at home. [Re: El Douche]
    #14065064 - 03/04/11 08:17 AM (12 years, 10 months ago)

Quote:

El Douche said:
I use rooting hormone for house plants. I just cut off a small branch, pluck the bottom leaves, dip the stem in hormone powder - including at least one nub (branching point), and place in tap water. Roots usually grow out in a week or 2. You could also place the dipped cutting in potting soil or other media and put a humidity tent over it untill the plant grows roots (you can use an inverted cut 2-liter as a tent).

I don't know how this will work with your specific plant, as my described method does not work on all of my plants.

You could put some Miracal Grow in the water.

I would think that agar would be difficult because it is prone to contaminants. The hormone used to come with fungicide, but the last time I looked it could not find the combination.

The hormone is toxic, so don't breath the dust or get it on your skin. A very small amount will kill an animal, and it is used in small amounts to kill fish in lakes.




Quick question here...what PH is suitable for this?

7 or so?  Sorry for the n00b question but I am interested in doing this to multiply some Pereskiopsis.  Thank you in advance!


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Re: Micropropagation at home. [Re: IHuntem2]
    #14065651 - 03/04/11 10:54 AM (12 years, 10 months ago)

A link I posted shows how to do it all for pereskiopsis with pictures... :laugh:
Its the amateur micropropagation/plant tissue culture thread at the laboratory.


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Re: Micro propagation at home. [Re: caricapapaya]
    #14065869 - 03/04/11 11:37 AM (12 years, 10 months ago)

Quote:

caricapapaya said:
Doc-

That looks good with no contams, the leaf pieces are still green (a good sign) as for the root segment....I dont see it looking like a shoot.





Not 'still' green, they started brown.
Root may or may not transform into a plant, that's what this experiment is about.


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Re: Micro propagation at home. [Re: caricapapaya]
    #14066211 - 03/04/11 01:02 PM (12 years, 10 months ago)

Quote:

caricapapaya said:
if a shoot starts to grow, it will typically be green (after a little while anyway, especially if exposed to light,




I cut up the second seedling on 3/2, I just moved one jar into my mushroom greenhouse.
It will get bright 12/12 light, but indirectly, the lid of the jar shields it.


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Re: Micro propagation at home. [Re: Doc_T]
    #14066276 - 03/04/11 01:17 PM (12 years, 10 months ago)

Keep it up. You never know what will work. I've seen some very weird stuff happen in a petri dish.

:thumbup:


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Re: Micro propagation at home. [Re: caricapapaya]
    #14066454 - 03/04/11 01:56 PM (12 years, 10 months ago)

Oh, no doubt. This is just my first try.
And it's something nobody else is even doing (yet), I'm surprised it's worked even this well.

So, if I'm working with leaf bits I want sucrose in the mix, but not if I'm trying to work with root material?
And what about dextrose? Useful or no?


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Re: Micro propagation at home. [Re: Doc_T]
    #14066634 - 03/04/11 02:27 PM (12 years, 10 months ago)

sucrose is the best for everything as far as I know - and probably the very best for roots, because that is how they receive their sugars - the leaves manufacture glucose at some stage and might be more apt to be able to use it.
Sucrose is just used for everything as far as I know..


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Re: Micro propagation at home. [Re: Burbles]
    #14068462 - 03/04/11 09:41 PM (12 years, 10 months ago)

yes, put sucrose in everything. I am sure there is a protocol for something that doesnt use it, but for the most part, use it.


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Re: Micro propagation at home. [Re: caricapapaya]
    #14072118 - 03/05/11 06:06 PM (12 years, 10 months ago)

Here is some info on various OTC sources of plant hormones...

http://www.thelaboratory.org/talk/index.php?topic=33.0
Including extracting gibberlic acid from seeds, and a few other good places..


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Re: Micro propagation at home. [Re: Burbles]
    #14073259 - 03/05/11 10:25 PM (12 years, 10 months ago)

You're coming at this from the wrong end.
We don't links to how to make hormones.

We need to know which hormones, if any, are required for this to work.
THEN we can learn how to make them. You're confusing data with information, you've been doing it all through this thread.
There's several people that have posted really irrelevant links.


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Invisiblecacharstar
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Re: Micro propagation at home. [Re: cacharstar]
    #14073331 - 03/05/11 10:42 PM (12 years, 10 months ago)

Quote:

cacharstar said:



I found this research done on the best adjustment of the ms medium for hemp but am having trouble understanding exactly how to carry out the mixing without a firm background in chem.. Perhaps we can help each other.:rasta: Here is the linkhttp://www.pakbs.org/pjbot/PDFs/41%282%29/PJB41%282%29603.pdf
.




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OfflineBurblesV
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Re: Micro propagation at home. [Re: Doc_T]
    #14074659 - 03/06/11 08:46 AM (12 years, 10 months ago)

Quote:

Doc_T said:
You're coming at this from the wrong end.
We don't links to how to make hormones.

We need to know which hormones, if any, are required for this to work.
THEN we can learn how to make them. You're confusing data with information, you've been doing it all through this thread.
There's several people that have posted really irrelevant links.





I thought the goal of this thread was to do micropropagation using OTC materials?
Of course it is going to need hormones... unless you are simply trying to grow a plant in agar... growing a plant in agar is easy - not so with getting the right ratio of certain types of plant hormones to form undifferentiated plant cells, or to cause them to grow shoots and roots...


What is the goal of this thread?
I was simply providing things that are useful for micropropagation in general.

What sort of data are you wanting too see, if any?


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OfflineDjcorbetto
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Re: Micro propagation at home. [Re: Burbles]
    #14090846 - 03/09/11 04:20 AM (12 years, 10 months ago)

I think he may be trying to say that all this extra information is confusing the thread. I read this to see how his tissue culture is going because im interested in tissue culturing cannabis plants, so to me all the extra bits are rather confusing, every time anyone asks about tissue culturing i send them to get the plants from test tubes books as thats the best thing you can have. I wanted to learn from this thread the method that doc t is using to see if it will work and efficiently produce some good cultures and the recipe that he's using because Ive heard of some weird things being used in the agar but never miracle grow. Keep up the good work Doc T, I've got my baby good jars coming in the next couple of days so i can start looking at the right ingredients that i need. Good luck


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Invisiblelucas_southoz
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Re: Micro propagation at home. [Re: Djcorbetto]
    #14107022 - 03/12/11 01:12 AM (12 years, 10 months ago)

Much happening with em doc?


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Offlinehumawebdesign
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Re: Micro propagation at home. [Re: lucas_southoz]
    #14107334 - 03/12/11 03:22 AM (12 years, 10 months ago)

i got some nice idea from all you peoples post hope my iea will work if everything goes fine then i will post it here.


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OfflineDjcorbetto
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Re: Micro propagation at home. [Re: humawebdesign]
    #14107381 - 03/12/11 03:51 AM (12 years, 10 months ago)

yo should tell us while you are doing it, that way people can give you advice on what to do as you are doing so you have less chance to fail and you might produce better results. of course set up a new thread for it


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Invisiblecacharstar
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Re: Micro propagation at home. [Re: humawebdesign]
    #14107487 - 03/12/11 04:50 AM (12 years, 10 months ago)

Quote:

humawebdesign said:
i got some nice idea from all you peoples post hope my iea will work if everything goes fine then i will post it here.





Good luck.  Post a link back here if you start a thread...


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InvisibleDoc_T
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Re: Micro propagation at home. [Re: cacharstar]
    #14135654 - 03/17/11 08:11 AM (12 years, 10 months ago)

First attempt: not successful.
The one bit of leaf seems to be doing ok, maybe even growing. But it's not exactly thriving.
Meanwhile, some sort of contamination has happened, most likely bacterial.



Next attempt: cacharstar supplied some cactus samples I'll be trying.
Growth medium this time is half-strength MG water, plus sucrose.
Updates in a couple weeks I hope.


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OfflineDjcorbetto
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Re: Micro propagation at home. [Re: Doc_T]
    #14137983 - 03/17/11 05:20 PM (12 years, 10 months ago)

I took delivery of baby food jars yesterday so im procuring my supplies and then i will start cannabis tissue culture, i will let you know how it goes.


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InvisibleDoc_T
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Re: Micro propagation at home. [Re: Djcorbetto]
    #14137996 - 03/17/11 05:21 PM (12 years, 10 months ago)

Woot!


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OfflineDjcorbetto
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Re: Micro propagation at home. [Re: Doc_T]
    #14142953 - 03/18/11 12:20 PM (12 years, 10 months ago)

Quote:

Doc_T said:
Woot!




My plan has been to tissue culture, rare strains of the cannabis plant, i was waiting for my friend to send me the baby food jars that he had collected for me. just need to buy the rest of the ingredients now.


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Offlinecaricapapaya
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Re: Micro propagation at home. [Re: Djcorbetto]
    #14143236 - 03/18/11 01:34 PM (12 years, 10 months ago)

yes, the contams are bacterial. I think you'd have better luck with microprop in the strict sense of multiplying by axillary buds.

I dont think you'd even need hormones.

let us know


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