Or_Grow's Tissue Culture Journal

The fruits of our labor. We welcome all types of plants, but grows posted here must be legal.
Or_Gro
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Day 24: Hopin to set up spare bathroom, as cleanroom, today; begin work on multiplication/replication stage tomorra....
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Will be cutting growing tips (circled areas, above) off of cuttings, and transferring them to agar media tubes/vials containing sugar (nutrient) and branching hormone.

Will run for a couple weeks and decide whether to swapout media to grow longer, or divide plants and transfer to rooting hormone...
Or_Gro
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Prep for Step 2: Multiplication/Replication

In the first step, i did the sterile work in a 4x4 tent, to demonstrate that these techniques can be done satisfactorily is a small space, using relatively inexpensive tools and supplies.

This time, i’m using a larger spare bathroom. It will provide more working space, and a good test of the capability of the hepa fan i’m using.

All non-essential materials were removed from the room; vents were closed; hepa filter/fan was positioned and turned on; and all working surfaces were cleaned, wiped down with 70% isopropyl alcohol, then sprayed with lysol.

Tote being used as a glove box was wiped down inside and out with 70% iso, then sprayed with lysol.

All tools and supplies not being sterilized in pressure cooker were wiped down with 70% iso, and placed on a sterile working surface.

Here are the media prep tools & supplies:
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2 tablespoons of sugar were mixed in 900mL of distilled water, along with the 2 fl oz of hormone, micronutes, and preservative and pH’d to above 5.8. Powdered agar was mixed in, and pH’d to 5.8.

20mL of solution was transferred to the flip-top plastic vials:
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Vials and other items to be sterilized were placed in pressure cooker:
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and sterilized at 250F and 15-20psi for 20 minutes, then cooled overnight.
Or_Gro
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Day 25: Gonna cut them up and transfer to mult/replic media this morning.

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Or_Gro
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Day 1, Multiplication/Replication Stage:
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Growing tips removed from stem
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Dying/damaged parts/extra tissue removed, leaves trimmed
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Growing tips inserted in agar-nute-hormone media
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Vial capped, wrapped, labeled
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Or_Gro
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So, i like the microclone kits, they come with most everything you need.

However, it would be useful if the directions included a short, straightforward set of steps on how to do the process with current materials in the kit....as opposed to current format that lumps theory and process together in a long document, based on older kit supplies.

Basically, some of the how-to is out-of-date and/or missing, and difficult to sort out from the theory.

I have some background, so i was able to muddle thru, but i can see how noobs might get lost.

A couple examples of things to clarify:
For stage 2, there are two diff sets of branching media, without any explanation of which to use, and when.

The vials have evidently changed to fliptops, there is no description of how they should be dealt with in the pressure cooker, if you close the lid, you risk them bursting open/spilling/boiling out; if you leave them open, you expose them to contamination when releasing pressure/removing lid.

One other improvement, would be having smaller “intro” kits, which provide the first time
user with the complete experience, but without the commercial-scale quantities of containers and media, which makes the kits expensive and wastes media that is not portioned for re-use, and provides way more containers than necessary to learn the process.

Or, portion-size the media supplies, maybe a 1L supply could be portioned into 4x250mL batches.

Finally, i understand the seller would like to sell every component at a profit, but simple things like the dichlor sanitizer could be easily sourced by user, if kit maker would just demo how to calc the amount needed of:
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In future, i’ll just buy the media mix, maybe some containers, source everything else myself...tho, i’d recco a complete kit for the first time doing this.
Last edited by Or_Gro on Mon Jul 08, 2019 6:07 pm, edited 2 times in total.
Or_Gro
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Day 2, Multiplication/Replication:
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Or_Gro
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Day 3: No obvious changes yet.
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damien50
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This is really awesome and something I could make room for.

Do you have a full supply list together of what you consider essential and or beneficial. I'm thinking of doing sog with 30-64 plants and would rather do this than maintain my several mother plants. Plus I wouldn't need to use my cloner lol.
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