Or_Gro’s Tissue Culture Journal

Or_Gro

Or_Gro

Well-Known Member
I’m interested in doing small-scale tissue culture of cannabis, primarily for gene banking and strain trading. I have a HEPA-filtered laminar-flow fan, and a makeshift cleanroom (aka spare bathroom).

My first attempt will be primarily to build my cleanroom & lab technique, by using a storebought “tissueponics” kit, that is basically an in vitro version of aerocloning.

Later, i plan on learning how to do in vitro replication and rooting processes.

Anyone who is serious about plant tissue culture is welcome to participate in this thread.

Here are some background and reference links:

https://www.cannabistech.com/articles/cannabis-tissue-culture-101/

https://www.cannabisbusinesstimes.com/article/a-primer-on-tissue-culture/

https://mycotopia.net/topic/50104-tissue-culture-protocol-for-mj/
 
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Or_Gro

Or_Gro

Well-Known Member
StickyBudHound said:
I've recently heard about this. Very cool stuff. Someday...
Click to expand...
I think you would get into this...i’ll figure it out.. you can see if it’s useful and whether you have space for it...you can diy a laminar flow setup to save $’s..., rest is just a small room/large closet/tent, a pressure cooker, and some small items and lysol/bleach/alcohol....

Can even be done without laminar flow/cleanroom, by building a glovebox....just a little less convenient....
 
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Or_Gro

Or_Gro

Well-Known Member
Imagine:

Having a testube or baby food jar or deli container in your refrigerator, with your favorite mother and her clones, waiting for their turn in the rotation;

EA48479A-E047-4C84-8E9C-E0EB790FA165.jpeg

Or

Sending to your growbuds across the country a tiny piece of green plant that can develop into numerous primo ganja plants...

1AC5131D-B682-4CF0-A69C-D3C16061914C.jpeg

And beautiful af...

08CB88F9-5679-455C-B3A2-440BFD0E847A.jpeg

CA772224-F35B-45A8-9EA9-D4204C219DE7.jpeg
 
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led1k

led1k

Well-Known Member
Or_Gro said:
Imagine:

Having a testube or baby food jar or deli container in your refrigerator, with your favorite mother and her clones, waiting for their turn in the rotation;

View attachment 4344297

Or

Sending to your growbuds across the country a tiny piece of green plant that can develop into numerous primo ganja plants...

View attachment 4344298

And beautiful af...

View attachment 4344300

View attachment 4344301
Click to expand...
Sending to your growbuds across the country a tiny piece of green plant that can develop into numerous primo ganja plants... Priceless!

Thanks for starting this. Alternatives maybe useful? Short videos walking thru use at bottom of clone link.
https://www.horticulturesource.com/tissue-culture-microclone-kit-p21366
https://www.horticulturesource.com/tissueponics-starter-kit-36-p27106

Have you seen any info on required psi for the autoclave part? I have an "Instant Pot" rather than a regular canning one so...
 
Or_Gro

Or_Gro

Well-Known Member
led1k said:
Sending to your growbuds across the country a tiny piece of green plant that can develop into numerous primo ganja plants... Priceless!

Thanks for starting this. Alternatives maybe useful? Short videos walking thru use at bottom of clone link.
https://www.horticulturesource.com/tissue-culture-microclone-kit-p21366
https://www.horticulturesource.com/tissueponics-starter-kit-36-p27106

Have you seen any info on required psi for the autoclave part? I have an "Instant Pot" rather than a regular canning one so...
Click to expand...
Nice tutorial vids on first one. i have ordered the first one, i have the second kit now.

Not yet....but, normal pressure cooker temps & pressures (15 psi, confirmed) do the job for mushroom culture work...i consider that as, if not more, demanding that this.
 
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Or_Gro

Or_Gro

Well-Known Member
Or_Gro said:
Nice tutorial vids on first one. i have ordered the first one, i have the second kit now.

Not yet....but, normal pressure cooker temps & pressures (15 psi, confirmed) do the job for mushroom culture work...i consider that as, if not more, demanding that this.
Click to expand...
Pressure cooker sterilizer: pressure is 15psi, 250F(121C), for 20 minutes.
 
Or_Gro

Or_Gro

Well-Known Member
Ok, so i’m gonna start with this kit:

https://www.horticulturesource.com/tissueponics-starter-kit-36-p27106

Then use some of these plants with this kit:

https://www.horticulturesource.com/tissue-culture-microclone-kit-p21366

The first kit comes with these directions:

3A91A7A6-8EAF-438E-B90A-8F5B37BC49E9.jpeg EA6B81BC-18FA-41E5-80F3-98411F4F8397.jpeg 44BAA784-EAC6-4E8F-BC29-138BC4CCF7A4.jpeg D6E4E76A-3F94-4B22-BFBC-5CEDA3B3937A.jpeg C5001FF1-6FE2-4ECB-AAF3-05FFEAFB1B81.jpeg

Videos of the process can be found here:

https://m.youtube.com/results?search_query=microclone+tissue+culture

Start with the vid titled:
8D7F3C74-E791-4CDF-9EB4-90BFE00AAE86.jpeg

I’ll buy some supplies tomorrow and get started as soon as i have some time.

https://m.youtube.com/user/microclone100
 
led1k

led1k

Well-Known Member
Or_Gro said:
Nice tutorial vids on first one. i have ordered the first one, i have the second kit now.

Not yet....but, normal pressure cooker temps & pressures (15 psi, confirmed) do the job for mushroom culture work...i consider that as, if not more, demanding that this.
Click to expand...
led1k said:
Really excellent stuff! Was thinking more about how wonderful to preserve a prized pheno without having to keep a full blown clone around... :clap::clap::clap:
Click to expand...
As soon as that thought settled in my mind... I ordered the first one too. 15psi pressure cooker on the way.
 
Or_Gro

Or_Gro

Well-Known Member
Re: small-scale tc, one of my overall goals is to identify a process that has acceptable survival levels with minimal investment in lab equipment.

From what i already know about in vitro culture work, the primary source of failure is infection, meaning, sterile technique is one of the critical elements to success.

In commercial labs, proper sterile technique involves disinfecting all samples, tools, chemicals, nutrients, and equipment that come into contact directly/indirectly with the sample processing area.

Most homegrown tc processors can’t afford the equipment, chems, and tools for this level of sterility.

I’ll begin with what is shown in the video, and if contamination levels get too high, i’ll prep materials in a pressure cooker and do transfers in a room with a hepa-filtered laminar flow fan.

I’ll be using 70% iso alcohol for surface and hand tool disinfection (70% is better disinfectant than 95%, cuz it contains more water which slows the procees of denaturing protiens and the slower process leads to more complete protein denaturing); and NaDCC (dichlor chlorine powder, aka pool shock) and polysorbate 20/tween 20 (surfactant for better chlorine contact) for plant material and container disinfection.

Based on the Youtube vids by Microclone/Bill Graham, the chlorine disinfectant needs to be 500ppm chlorine (1mg/L=1ppm)...the following articles provide background and a methodology for making this solution (different sources of chlorine have diff chlorine content; e.g. laundry bleach is typically around 5-6% chlorine, the dichlor powder i’m using is 58.2% chlorine).

https://www.biotopeone.com/nadcc-pool-chemical-as-an-explant-sterilization-tool/

https://phc.amedd.army.mil/PHC Resource Library/TIP_No_13-034-1114_Prepare_Measure_High_Chlorine_Solutions.pdf

For mixing the various nutrient/hormone solutions, i’ll use boiled reverse osmosis water (since i have r/o water), but boiled distilled water is an alternative. Cover boiling vessel and bring water to a roiling boil, continue boiling for 3-5 minutes.

Water treated with chlorine can also work.

https://www.epa.gov/ground-water-and-drinking-water/emergency-disinfection-drinking-water
 
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Or_Gro

Or_Gro

Well-Known Member
If you want a very sterile environment, you will need a hepa-filtered laminar flow fan.

Here is a great source, https://fungi.com/apps/omega-search/?type=product&q=Laminar*+flow*+hoods*#type=product&q=Laminar* flow* hoods*.

But, they’re friggin expensive....

Here’s the diy version:


Detail mentioned in Part 2:

Published on Oct 12, 2018
Freshcsp Mushrooms
GET FULL BUILD PLANS: http://freshcap.link/diy-hood-plans

The Homestud Jeff helped TONS with this build, and has awesome content your should check out. YouTube: https://goo.gl/J4qvxW Website: http://thehomestud.com/ Instagram: https://www.instagram.com/thehomestud/
 
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