micropropagation? storage life of M and S Basal med with vit ???

[Dreamwalker.]

"To start learning your tech I'd recommend getting:
10 @ 1L Murashige and Skoog Basal medium with vitamins ( or share a 10L purchase amongst friends )
Agar or other naturally-derived gelling medium ( we'll skip the synthetics for now )
Hormones- BAP, IAA, IBA and Kinetin. You can buy these already made up in solution, they last three months in solution correctly stored so note the date you received them on the label
Coconut water, 100ml should be more than enough"

So I want some BAP for pupping, but I want to also start micropropagation (realistically it may be many months before I can have a lab/gear space, set up, and ready to go).

But as I want to induce pupping now and so order some BAP, It seems like a good idea to order

NAA & Murashige and Skoog Basal medium with vitamins- because the courier fee is high (toxic substance), best to just make limited orders, 1 courier cost..

I have read that NAA and BAP dry crystals store ok for up to 4 years

but I guess Murashige and Skoog Basal medium with vitamins as its sold by the L has a more limited shelf life.

Does anyone know what its storage life is likely to be?

and can I get by (beginning) with just NAA and BAP?

Agar I have.

Edited July 12, 2014 by Dreamwalker

[Darklight]

Kept as a powder solution in a sealed, dry container in the fridge or freezer, most TC media like MS will remain functional for many years. If it's in solution, whack it in the freezer and allow to thaw at room temp or slightly higher before each use. Don't microwave the frozen media in the bottle

I wouldn't use years-old media for research purposes in a research environment, but for home use in the early stages you won't even notice any physical changes in an unopened media sachet for a couple of years- the decay is not rapid if it is stored as a powder according to directions on the MSDS

As a beginner, you are going to have more of a challenge learning to keep your cultures sterile than you will have concerns about the age of your TC media. Focus on the former, not the latter, for the time being

You can certainly get by with just NAA and BAP, I'd get some IBA as well tho if you can, for general use.

Think about the project/s you have in mind while you place the order.

If you have a particular species in mind you want to work with tho, google to see what protocols are out there for your species and if there is more than one protocol, note which hormones they use and order those

If you are starting with a limited amount of parent material, that will dictate how many experiments you can run before you run out of starting material. However if you're working with something with an endless supply of starting material, like African violets, you will have different operational parameters

Even having only two hormones, NAA + BAP, and a defined media like MS, will give you more than enough variables to run an infinite series of experiments. Different amounts of each hormone, different ratios between each hormone, different media strengths and carbon source ( usually sugar ) amounts - you can run some really rigorous trials easily that will give you some great ideas about what to do next

My rule of thumb: start absolutely no more than 5 experiments at once. Experiments are like puppies, they grow up and have more puppies. If you don't watch them all carefully there are eventually so many of them that they become unruly and you will want to leave home to avoid them and the whole exercise will rapidly become pointless

Two or three experiments, run really well, recorded meticulously ( even if you make mistakes, just note them, they happen ) are worth 10000x more than something you started but didn't log properly

[Dreamwalker.]

Thanks DL...........I would never have considered freezing..great idea.

Probably using tissue culture for cacti...then maybe (giant bamboo, banana)

so maybe just M&S + vit

BAP

IBA (instead of NAA)

What do you used as a good source of distilled water? (I'm guessing auto distilled battery water or well filtered sterilised rainwater, may not be up standard)?

Edited July 13, 2014 by Dreamwalker

[Darklight]

What do you used as a good source of distilled water? (I'm guessing auto distilled battery water or well filtered sterilised rainwater, may not be up standard)?

Bugger, missed this, sorry

For pure research work on new species I use water from the supermarket that's sold for irons and stuff, prolly the same as battery water. If I have access to a good lab I'll grab water from a Millipore unit, 0.22uM, but it's overkill for small scale use. Supermarket ironing water is fine AFAIK , and I've had chemists tell me they use it for TLC and spectro as well if they're away from facilities

Once I'm comfortable with the protocol and I have spares for replication, I move to rainwater from my tank, filtered with a domestic water filter. I know it varies from season to season ( I checked- had it analysed years back over a few seasons ). It's never been a problem, but then I never dump all my work into a batch which contains a new variable, I always keep spares for later

So long as you don't use filtered rain or tap water for your initial work, you should be good

[Dreamwalker.]

thanks ...no worries...I got all the hormones in the fridge & the rest of the kit....just waiting on a window of time to get started.........grafting is taking most of my spare time at the mo.......so need to move up a level & get the tc. going....probably as Autumn approaches.....thanks for your guidance........

[Darklight]

No wuckas, sry to have missed your questions

Spring and Summer are best months for TC as plants are more actively growing which helps outpace contamination. Pref to start most species when not in flower. But if you have to choose... ;)

[shonman]

Very informative, Thanks!

I was starting out, got lots of hormones for plants.

Then they got old in the fridge while I worked out sterile technique and plain explant division!

Kept me busy almost a whole year!

When I went to finally use hormone to multiply in this case Mitragyna Speciosa,

which is ok where I am....

The hormone didn't work by the time I wanted to finally use it!!!

I had thought I was being thorough and was all stocked up with various hormones,

a lot of which are old too now.

So, I ordered more hormones from Carol @ Kitchen Culture Kits

who is a great source of info as well

In powdered form, these definatley store well for a longer time.

They are combined with solvent then diluted when ready to use.

With some varieties of warmer climate plants,

merely cooling them to a lower temperature will make then practically stop growing.

A temperature they consider winter.

Of course, don't experiment with 100 plants at a time, you may lose them in some cases!!

Thanks again Darklight for sharing info with everyone!!

BTW- Carol has developed some excellent tech for medical Cannabis, legal where she is,

Non medical is also legal there now, so she is following the rules.

Hopefully we will all be able to experiment with these plants legally in the near future!

There are many benefits to science, art, medicine and culture .

(edited out rambling philosophical digressions)

Edited December 28, 2014 by shonman

[Darklight]

Ta for reminder Shonman

Rule of thumb is hormones in solution store no longer than 3 months, pref frozen and thawed prior to dispensing in room temp water. Some store longer, but I wouldn't risk it at research phase

He's right tho, don't buy them til you have sterile cultures if you are just starting out

[Darklight]

Hey Carol's doing legal pot now? Lucky woman. I originally started TC so I could play with it but got distracted early on by kratom. Never got around to it

Prolly just as well, kratom was easier. Shame it's not legal any more, it's an excellent beginner's plant

[shonman]

I use distilled water from the store, too.

Feel guilty about all the plastic containers,used,

but I do take them to recycling.

I am considering gertting or making a water distiller,

however, when I think about how cheap it is to just buy the water

I usually go do something else and buy water when needed.

Perhaps a self distilling solar water distiller would be ok

just fill and do something else.

This could be created from a keg with the top cut off

and a few nice large stainless steel salad bowls.

Basicly, water in keg heats in sun

evaporates,

condenses on large salad bowl on top of keg

which is fillled with cool water or ice,

drips into smaller salad bowl,

inside keg on a stand.

stand sits in regular water, on bottom of keg

Small salad bowl fills with distilled water

later to be sterilized further before use.

(edited typos)

Edited December 28, 2014 by shonman

[shonman]

Carol is in the process of making a kit to sell for Cannabis micropropagation,

where legal of course.

Which may be in more locations over the next few years!

She has much wisdom to share, as does Darklight, regarding various kinds of micropropagation techs.

I would post what she sent me, but its probably better if you ask her yourself.

A search online will reveal her interesting ghetto micropropagation techniques,,

as well as a way to contact her if needed.

[Darklight]

I use distilled water from the store, too.

Feel guilty about all the plastic containers,used,

but I do take them to recycling.

I am considering gertting or making a water distiller,

Just get the water from the shops unless you're using more than 10L every day. Seriously. I wasted money on double distillation water units years back, the situation changed and I've never switched them on Think maintenance, think seals, think benchspace, plumbing, power

I've heard Reverse Osmosis units can be cheap and provide good quality water, just make sure you get one with easy to find cartridges. But really, stick with ironing/ battery water. Spend your $$ on other toys

Carol is in the process of making a kit to sell for Cannabis micropropagation,

where legal of course.

Which may be in more locations over the next few years!

She has much wisdom to share, as does Darklight, regarding various kinds of micropropagation techs.

I would post what she sent me, but its probably better if you ask her yourself.

A search online will reveal her interesting ghetto micropropagation techniques,,

as well as a way to contact her if needed.

Haven't corresponded with her in years, she prolly wouldn't remember me but her expertise was invaluable in my early days

Cannabis kits, hey? That should get a bit more interest in the field

Ta, no need to share private corro, but I should check her ghetto teks again, there have been some amazing advancements in the field of late

[shonman]

Kitchen Culture Kits is her company, for those who would seek information from Carol.

She is located in Washington State, USA

[Darklight]

Hey those newsletters from http://www.hometissueculture.org/newsletters.html are seriously interesting. Really good basic info. And free. Much stuff there from Australians too. Ta Shonman!

[modern.shaman]

May not be applicable to you but I use 'distilled' water produced from my dehumidifier for my TC. I bought powder forms for all my material and they had expiration dates on the containers assuming you store them according to the label. Most including MS are listed as lasting 2 years but I'm sure they can be used for longer, NAA is listed as only 1 year though.

You should consider getting PPM or an alternative. I've had great success introducing material however quick callous growth and pupping have eluded me so far. I'm only 3-4 months into it and am not giving up do to feeling close to making progress. I may transfer to a media without PPM to what I have last longer.

I've been working with cacti and will likely start a leafy plant just for faster results.

Edited December 29, 2014 by modern.shaman

[shonman]

I also consider PPM almost indispensable,

although my cultures have done well without it too.

One thing I did notice, which Darklight also pointed out elsewhere,

is that after several weeks in a culture,

the PPM seems to get used up.

Plants that were fine, sometimes show contamination after a while.

I am surprised at how I have recently kept about 400 explants

in the same medium

for about nine weeks!!

They are mostly all fine, with a few casualties here and there that never started in the first place.

[shonman]

I had not considered water from the dehumidifier,

that seems like a great idea, unless metals somehow find their way in from the coils where water condenses.