X-Ray Mutagenesis

I became interested in mutagenesis after reading about the Optimara EverFloris Space Violets program and eventually finding my way to the Wikipedia:Mutation_breeding page on May 27th 2019 where I wrote "might try this" on dobbs.town.

I began by trying to acquire a chemical mutagen EMS, with plans to dispose of it with a NaOH solution, but gave up after one attempt and the order being denied. Probably for the best! I continued to scour the internet for stories of people attempting to mutate seeds or something like yeast at home. I found someone who would bring seeds to their dentist to have them x-ray seeds as well as someone considering usage of UV-C light to induce mutations.

I knew I had read a way to use old vacuum tubes hooked to high voltage power supplies to create dangerous x-rays. Using the power of the Internet, searching, deal hounding, researching and designing (still in progress) I've been able to gather enough parts to have a "Mutagen Chamber" ready in the next few days. The last thing I needed was lead shielding. I did a quick test run with one of the 2x2a rectifier tubes and a cheap HV stun gun module ordered from a company in China off eBay. The Geiger counter alarm went off signalling over 100cpm and I switched the power supply off. It was a variable power supply so I wasn't even using the full HV output, I was slowly turning it up listening to the chirps steadily increase in frequency of occurrence until the alarm started beeping.

I found a listing for 10x 2x2a tubes on ebay for $26 shipped, making it $2.60 per tube. The HV taser modules (I ordered 2 since I've heard they have a tendency to burn out) were bought for about $2 each. The control circuit is still being designed, I need to figure out better isolation between the microcontroller and the HV module, there's just too much noise, or maybe I need to figure out a more robust timing circuit because I want to run pulses where it's on for 2 seconds off for 2, something like that so the tube or HV module doesn't overheat. The lead is 33 pounds of 1/16" lead sheet roll, was able to get that shipped for a total of $50, the priciest thing so far.

I used FreeCAD to design and 3D printed a socket and "germplasm ring", as well as a cathode electrode connector so I can simply jam the HV module wires into the plastic connectors and start getting x-rays.

Hope to get more of this down and organized. I've got a first generation of 'Wisconsin Fast Plant' seed as well. I want to get a petite population going so I can grow a lot of plants dense and check for mutations. Would be AMAZING to get some viable morphological changes.

tube-holder

8/14/2019

I've got my hands on some 1/16" scrap lead sheet, I had to soak it in water to get some paper off of it, the glue dissolved easily.

rolling cutting sheet prototype

The "welding" I attempted with the propane torch didn't go so well, I'm actually thinking of securing everything with duct tape... should work! I also want to design a detachable ring that I put onto the lid and lower into the chamber to make the fiddly loading/extracting of the seeds easier.


9/6/2019

I put the lead container together using duct tape and little nails to hold everything. It ended up sitting around for a while while I considered what I was doing (or whatever) up until today, where I decided I was going forward with this. I clipped an old FM antenna for the wire to hook up the HV module to the variable power supply. I selected one of the large pins at random on the 2x2a (there are 2 sets of pins, 2 small and 2 large) basically following Dangerous Laboratiories page on DIY X-rays.

2x2a

Tube Data

and hooked that one up to the end of the HV module that seemed to glow with more corona than the other (might not be correct here...), then the other HV line to the tube's anode on the top.

While exposed directly to the energized tube (about 10cm away) the counter instantly peaked (>600cpm), I found leakage of about 54cpm close to the lid where it wasn't sealed. After getting the active readings, I ended up putting my left over Wisconsin Fast Plant confetti seeds (about 25 of them) into the chamber. I 'cooked' them for about 4.5-5minutes with 3vdc going into the HV module. So I'll be planting some of my 2nd generation fast plants, unexposed first gen seeds and these x-ray exposed seeds later today.

xray1 xray2 sed

I have the seeds planted in 4 rows: A, B, C, D with columns 1-16

  • Row D 2-15 X-Ray Exposed
  • Row C 2-15 F1 Generation of purple stem WFP
  • Row B 2-15 Un-exposed Confetti Seeds
  • Row A 2-14 X-Ray Exposed

9/8/2019 7:40am

plant-matrix

Germinated:

  • Row A: 2, 7, 9, 12, 13, 14
  • Row B: 2, 9, 14
  • Row C: None
  • Row D: 4, 9, 10, 11, 13, 15

Interesting that the exposed seeds are germinating so quickly compared to the rest. The F1 seeds are not the most vital since they spent half their life in low light conditions (probably).

3:15pm

Ok, so I did some more experiments with the x-ray stuff and I think the x-rays I was producing at 3vdc were too soft for the seeds to be so viable. They should actually be taking MORE time (because of the time required to repair the DNA). I turned up the voltage to ~4.5v and was able to get the geiger counter going wild outside the lead container.

So I exposed my collection of cat's claw seeds that I collected while running through an alleyway. After about 5 minutes at 4.5vdc into the HV module + 2x2a I spread the seeds through a little section of the garden. I've read they can take anywhere from 3weeks to 3+months to germinate, I've had a small pot going with no success yet. We'll see what we can pull from the chaos!


9/17/2019

tricot small downward flowers stem cotyledons
  1. Looks like there's three cotyledon leaves on that first picture (A7)
  2. Both cotyledon leaves appeared diminished, but secondary leaves still grew, although small(A9)
  3. The 2nd to last picture shows a plant that have given up radial symmetry, and the flower buds appear to be pointing down(A14)
  4. And finally the last picture is one of the most interesting mutants so far, the cotyledon leaves have extended stems (D8)

Buds have begun to form, there is a lot of variation with the height of the plants which makes lighting difficult.

Currently Growing:

  • Row A: 2, 6, 7, 9, 12, 13, 14
  • Row B: 2, 5, 6, 8, 12
  • Row C: None
  • Row D: 4, 8, 13, 15

None of my F1 seeds germinated :( (Row C). The x-ray exposed rows (A and D) showed at 40% survival rate (11/27 seeds). The unexposed seeds showed at 35% survival rate, which is odd because the first time I grew M0 fast plant seeds they had almost 100% survival rate out of 42 seeds. I suspect there was a fungal infection of some sort carried over from the first round of seeds combined with over-watering. I need to come up with a better watering solution.


9/19/2019

I exposed some Datura pollen to x-rays, 3vdc for 5 minutes like the seeds. I'm afraid the higher voltage may sterilize them if I ran it for that long. Might be better to just run it at a higher voltage for just a few seconds. Hopefully I'll be able to figure that out by plotting survival rate vs voltage and time.

I used a small pollination brush to collect and pollinate a bunch of flowers, maybe 8 in total, all marked with rings cut from a bobba straw and slit to allow it to be wrapped around the stem. I'll have to have a special area to try and germinate seeds from any pods that form from those flowers.


9/26/2019

A15-mold D15-silique A13-second-flowers A13-full A7-flower-explosion

Pictures from left to right:

  1. A picture of the mold that has taken over the whole setup, it was there the first time, but it's come around much sooner and much stronger than the first time. I need to probably just toss this setup and think of something new.
  2. Picture of D15's deformed seed pods or Silique not sure what their final form will be, but the initial plant wasn't too different besides having one shrunk cotyledon.
  3. A13 produced a 2nd set of flowers from the first Y branching after the first set went immediately from bud to silique production seeming to skip pollen production and the flowers even seemed to die off before even existing.
  4. A full shot of A13
  5. A7 was the one with the tri-cotyledon and grew a stem with weak flowers all the way up to a bunch of flowers that seem to be producing pollen and I've been pollinating it by hand and keeping the pollen sticks labelled and separate.

10/4/2019

A13-roots A14-buds
  1. A13 roots since they stood out to me today. I still have the light on and am letting the seeds mature, hopefully some will be viable.
  2. A14 cluster of compressed buds, no attempts at flowering just yet