Genesis of the mutants - or shuffle my base pairs please!

[ian]

I've been breeding up slime strains for a while now, hoping for chance and luck to conspire and give me something different. Ok, I got Daywalker which is a very handy little mutant but that's one out of hundreds of trials. Waiting for nature to give me my mutants is taking way too long, so I think it's about time I gave biology a little shove in the right direction and gave our physarum's genome a little tickle. 

Making random changes to an organisms DNA is easy - and has been a stalwart of plant breeding since the 1940's, pretty much all of of our modern agricultural plants have their ancestry in atomic gardening. ( https://interestingengineering.com/atomic-gardens-how-plants-were-b... ). 

Of all the methods I could use to make mutants, radiation makes the most sense - mutagenic chemicals probably don't belong on the kitchen ( although caffeine is a mutagen...), I don't much fancy getting myself a cobalt 60 source ( though I could lay my hands one, I think the physics dept might notice it missing ) - gamma rays might just tweak my own DNA too and not in the Bruce Banner direction - But I do have a source of mutagenic radiation that could fit the bill - UV.

I've got an aquarium sterilising lamp which makes light around the 250nm mark - right where DNA absorbs the most. They sterilise by ionising molecules, breaking apart bonds in the DNA and forming new ones that really shouldn't be there. Blast your sample enough and the DNA just falls apart - but that's not what we're after.

We want to half kill things - literally in fact - zap cultures until about half the cells die. The remaining cells will have catastrophic changes to their genome and most will shortly die but in a few DNA repair enzymes then attempt to fix the damage, often inserting the wrong base or skipping a section - in other words, makings mutants that hopefully look or behave differently. 

The UV lamp is my safest option - the radiation goes away when you turn it off, but it's not a safe option - these lamps burn skin very quickly, cause cataracts and cancer, job 1 on my to do list is make an enclosure that's light proof. 

Watch this space....

[ian]

I'm working with the haploid amoeba phase here for a couple of reasons - firstly, they only have one copy of the genome per cell which means they can't repair DNA by splicing from the other set of chromosomes ( so higher mutation rate ), they also only have one nucleus per cell unlike the plasmodium so there's not going to be back up for damaged nucleii. Lastly, physarum is yellow for a reason - they absorb blue/uv light very strongly - their pigments are an effective sunscreen, 20mins under the lamp didn't kill a Mazie.

Spores are easy to germinate, they need to have been dry for at least 3-4wks, mine are a year old, I just put a spatula's worth of spores into 10ml water, added a few finely powdered oats, mixed it and left it for a week. Each petri gets 15drops which is about 250 microlitres. The agar is 2% food grade agar in water, not sterilised, no nutrients. 

I spent a fair bit sketching out wooden box designs before I realised I was being stupid - yes the UV lamp is dangerous but it's also not going to be running for long so I went with this - 

Yep, a polystyrene box with the bulb suspended by garden wire. - it does the job and the fluorescent glow emanating from the box reminds me it's on. By the way this kicks out a huge amount of ozone - sore throat levels so if you're playing along, please ventilate the room. 

The box in the bottom is just to raise up the dishes a bit - with the agar in there it's 10cm from bulb to petri dish surface. 

Exposure time - pure guesswork here, in the lab 20mins is enough to sterilise anything but our bulbs have a bit more oomph. I need to find the exposure time that kills about half of the amoebae present - but without counting the amoebae. I do have cell counting kit at home but I'm trying to come up with methods that need the minimum of specialist kit so the plan here is to just count plasmodia as they form and use the colonies from the highest dose plate that still gave colonies. 

There's space to fit three petris under the lamp so I've done 5 control dishes then three each of 5,10,15,20mins. They're incubating at 20oC, I'll give them a few days then look for signs of amoeba foraging but it'll be a min 3 weeks before plasmodia form,.

[Julian Northey]

Hi Ian,

Please keep us posted on the progress of your mutagenesis and forward genetics experiment/s.

[Skadi]

Haha . "Damage the fuck out of them so they mutate faster" is my kind of logic .

Right now i·m working on a similar project , whereas in mine i·m using meltwater from heavily polluted snow to create an environment swimming with bitumen . It looks pretty dead in there , yet there·s lots of air pockets showing fermentation , so ,
s o m e t h i n g is growing in there .

[Patrick Barry]

How did this go? Anything interesting emerge?

[ian]

Sadly nowt - even the controls. Trouble is I've given up my study to a guest long term so repeating is a pain but I have a plan I've just made more spores, they need a few weeks dormancy then I'll incubate them behind the EM power supply in the lab - it's warm and no one goes near it ;)