Liquid Culture Vials - Un colonised

[redsmurf]

I have seen around that there are culture vials that are not yet colonised or inoculated.

does any one know where to get these in Oz as i am not sure that they would not be siezed by customs.

had a short but by no means an exaustive look. think they may be the solution to summer hiatiaus periods without complete restarts

[goneski]

Culture vials? Do you mean test tubes used as culture slants?

[satyr]

An alternative would be "Master Foods" herb jars. PC-able, and work well for long-term storage in fridge.

[redsmurf]

yes i thought about these as a fall back i also looked at different test tube arrangments but need to do more research into media.

[goneski]

Are you wanting long term storage to build a culture library? If so, agar in master culture slants seems to be the way to go.

Short term storage, you can use agar in jars or PP5 containers. Or even liquid culture.. Really depends on what you're wanting to achieve.

I've just been working with liquid cultures in baby bottles, nipple inverted and filled with silicone as an injection port.. and a glass shard in the bottom to break up the mycelium.

I haven't refrigerated them yet, but they're getting to that stage (the growth really needs to slow down / stop, as I've recently been having problems drawing up mycelium via a 16G needle,

even after shaking the bottles and breaking up the mycelium). It seems quite common for people to use LC that's 1-2 years (or more) old, so that's possibly an option.

Paul Stamet's books are a good starting point. To be honest, I haven't read through them in their entirety or even tried 10% of what he writes, but

they're a treasure trove and should answer your questions.

I tend to just go with teks on the usual sites, and when I need a definitive answer, go straight to Stamet's books.

Edited January 15, 2013 by SYNeR

[Cue]

Sounds like you need to move up to a 14g needle SYNer.

I have used LCs that were up to 6 months old, but I like to use them within 3 months though. But I store mine in a cool closet. I would store them in a fridge if I had room for a small one that I could dedicate to mycology. I'm just too paranoid about putting them in one that is used for food. There are just too many micro-organisms in one used for food storage to allow me to feel comfortable about putting a culture in there.

Times may vary depending on species of course.

Edited January 15, 2013 by Cue

[goneski]

Sweet, so 14G needles work fine?

I've noticed even with the 16G, when injecting through silicone, you tend to get a chunk of it in the needle.. So I wasn't sure if it would be an issue using 14G.

Now I just need to find some. I've only ever come across 14G piercing needles (no hub to connect to a syringe), or blunt 14G needles

Edited January 16, 2013 by SYNeR

[redsmurf]

may of gotten my wires crossed a bit but i am talking about making sterile live culture vials that can be refridgerated and used as an intermediatry stage between spore syringe and inoculation.

kind of like a mini inoculation but one that can be put on hold but not as long term as slants.

I have seen the vials for sale online but am worried that they would fall foul of customs dispite containing no viable organic matter.

so i looked into it a bit and its easy enough to get the vials sterile and empty now i need to look into what to use to make the liquid culture, seen many variations.

hope that makes sense

[trucha]

14 gauge do chew up the septums. While they are still in the hood try smearing a little RTV silicon over it afterwards to be sure its resealed.

Check the large animal care section at your local farm supply or ask a vet about getting some fat needles. 16s are easy to find but 14s are available with a little looking.

Or buy a jerky maker's "flavor injector" kit for just its 'needle'. I don't know its size but its bigger than 14 and has a luerlok fitting.

Not everything likes being broken up enough to go through a syringe. I found both shiitake and hericiums prefer me to use sterilized and then flamed forceps to pull loose pieces of the mycelium in the culture. Establishment and growth after inoculation have gone far faster for me with those particular mushrooms.

For what is being described LC in a small mason jar kept in a refrigerator would work for many months. Or at least it has been doing fine for me.

Edited January 17, 2013 by trucha

[trucha]

Munson's ME or light agave nectar has given me the best results. Agave produced slower growth than any other media I've tried but the colonies have always been pristine so I like it. (I mostly use LC in the process of cleaning up wild clones.)

Honey or Karo promoted growth the best for me but showed the most contamination as well.

[goneski]

may of gotten my wires crossed a bit but i am talking about making sterile live culture vials that can be refridgerated and used as an intermediatry stage between spore syringe and inoculation.

kind of like a mini inoculation but one that can be put on hold but not as long term as slants.

This is what I use baby bottles for. Turn the nipple upside down, and fill with red RTV silicone (I think almost any silicone will work as long as the temp is rated high enough for the pressure cooker).

Mix up a solution of 4% honey water (use distilled or spring water) and simmer it for 10 - 15 minutes. Pour into baby bottle, replace lid and tighten lid, then untighten a little so it doesn't seal and create a vacuum

when PCed.. Pressure cook for 10-15 minutes. Once cooled, tighten the lid again.

Done.

Now, you can inject spores through the silicone, and you'll have a live, multi-spore Liquid Culture grow out. Store in the fridge when it's grown out sufficiently to stall the growth. When you want to use some, draw up the solution into

a syringe, and inoculate grain jars or whatever.

Instead of shooting spores into the baby bottle, you can also clone a tissue sample from a fruit and drop it in and it will grow out. I did this a couple of weeks ago and inoculated some grain jars 14 days ago.. They should have

reached 100% colonisation by this afternoon.

You can then keep your baby bottle as a master if you want, and then propagate 2-3 cc to more baby bottles to keep the culture going.

You can waste time and money on finding/buying proper vials, but I find the baby bottles extremely convenient.. $3 for a pack of three. Just make sure you get the short, stubby ones so it's easier to draw out the solution.

I tried a 4% Karo solution, but compared to honey I found the growth really slow and stunted. LC growth in honey is pretty awesome. I do have problems with sediments from time to time, and the solution isn't crystal clear, but eh,

you're going to have to grow it out on grains or whatever first anyway to confirm that it's contam free.

[trucha]

Another nice thing about that baby bottle approach is the lids aren't going to eventually get rusty like with a canning jar.

[endorfinder]

Hah, I'll sell you some LC jars, presterilised with injection ports if you're truly that lazy

But really based on your question you need to read up a bit. Going straight to liquid culture is a less than ideal option since you need magnetic stirring to be remotely efficient about it, and it makes spotting contamination next to impossible. You're better off going to agar -> LC or perhaps even with grain liquid culture (GLC) depending on your situation.

Not sure about the source rules, but there is an Australian vendor of mushroom supplies that carries the bulk of what you need, if you can't find it locally. As well as the usual staples, they sell things like rubber injection ports and filter disks that make assembling idiot-proof containers easy. They also have agar starter kits with petrie dishes and such...

It's not hard to do things on the cheap from honey/corn syrup/malt extract, agar powder from the asian supermarket, some glass/PP (type 5 plastic) containers and a way to sterilise said containers (pressure cooker preferably). You may already have the things lying around the house.

[Bigred]

i use the urine jar's 50 cents each from my chemist they last in the pressure cooker

and inoculate in front of my flow hood ,but i just replace the silicone each time 16

gauge work fine for me just sharpen it on a match box when it gets dull

i had a blood test the other day and at the pathology they had these awesome

vials filled my pockets was thinking they will be great for storing strains

but im thinking clamp connections fascinating subject

[goneski]

urine jars.. awesome idea

[Bigred]

get the ones with the yellow lids they are pp5

[endorfinder]

@bigred the BD Vacutainers are actually pretty handy... guaranteed sterile.

[Bigred]

i know thats why i want to use them for cold storage and clamp connections

would also be good to send spore's as the syringe is classed as paraphernalia

[endorfinder]

i was thinking about using hot candle wax to seal barrels so you don't have to send sharps... anyone tried anything similar?

[Bigred]

candle wax will be to brittle the urine jars are sterile if un opened so

if your working under a flow hood i cant see why you cant just put

it in there

[endorfinder]

good point, even an eppendorf tube would work if you have concentrated solution.