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Tissue Culture Cloning

Agar is widely available from Amazon
https://smile.amazon.com/s/ref=nb_sb_noss_2?url=search-alias=aps&field-keywords=agar, or scientific or lab distributors.

If that isn't an option, a college science department, health clinic or veterinarian might be willing to order it for you, or if they're in a good mood, might give you a big scoop of it for a couple bucks. Might even come in little petri dishes.

I also watched a few of those NPK university videos, VERY detailed. I forget which one it was, but the guy did go over hormones, their roles and different available products like NAA, oligosaccharides, auxin, etc. I think he was talking about cloning, but still, I would assume the same hormones are still triggering the same response from the meristemic cells. If there are any other hormones, I'm sure you can replenish all the supplies a lot more cheeply than buying another kit.
 
Hi Greenthumb

Ok checked up your link to find out about Magenta vessels. That is just a brand name for a polycarbonate container available from any lab supply company. The other kit although more confusing is a lot better kit than the first one you mentioned. at least when you have used the media and hormones you can replace them.

The bit about needing an oxygen supply is totally wrong and is very dangerous. If you see a website suggesting this then I would leave the site and disregard everything they said. A typical way of keeping an aseptic work area is by the use of a naked flame inside your laminar air flow cabinet and every time you move from working from one culture to another is to dip your scalpel or forceps into 80% ethanol and then place over a flame to resterilise them. Open flames and oxygen are extremely dangerous, so unless you are using 2 separate work areas you run the chance of blowing yourself and the lab up.

You mentioned that you were confused about the reference to hormones. I had assumed that you had a general knowledge about what they were and how and why they are used. I hope you don't expect to just buy a kit and start growing. Buying what you need is a simple matter but knowing how and why you use them, is the complicated part. What one type of plant requires to grow successfully in vitro may be completely different for another plant. Even using different starting material ( leaf, Meristem pieces ) from the same plant will need different types and concentrations of hormones to grow.

I suggest you look at phytotechlab.com and look at their technical page as a good source of information. They are a very good company and what you read on that site is reliable and correct. It would be impossible for me to give you, step by step instructions without knowing exactly what type of plants and explant material you want to work with. So my first suggestion is to work out exactly what you want to do.
 
That's why I said that kit was not very good as they don't tell you what the different components in their media is and unless you know that, you simply cannot replace what you have used.
 
hey Greenthumb , i really REALLY admire your attitude and your confidence and hope all of this and more happens for you...dudn't sound like you will be but just wanted to echo your sentiment of ....don't be deterred by people telling you you can't or you shouldn't....
 
if there is *ONE* benefit to having this particular plant being precluded from serious, double-blind, peer-reviewed, the whole ball o'wax, so to speak, STUDY by the scientific community it might be this^^^ that whatever works works.... and there is no "governing body" to tell you, the individual "scientist", otherwise.
 
Well a word of advice if you ever need someone to come plant sit... DON'T EVER ASK ANYONE!!! I will spare you what happened to me, but just sum it up to say it cost me 5-years scrubbing mold. The 2nd time I came home to NOTHING, my "friend" of many long years took even the curtains and toilet paper along with everything else and then to my disbelief couldn't even figure out why all the plants died and why he couldn't make it work since he had all my stuff, and knew everything I told him to do for them.
 
Oh yeah I know the video's over there very well, Harley loves educating people about hydroponics and plants... I have spoken to him a few times in email and sat in on the classes, which after the video ends, then he says "Does anyone here have any questions". He takes all the time he has to answer everyone's questions before closing the My Meeting window where people sit in on his classes for Free. I have also asked him once about these kits and that is when he told me the book he is reading as he himself as he is planning on putting together a class on the subject of Plant Leaf Tissue Culture, but then I heard he is involved in a study again right now so that work is on the back burner. But I will keep waiting for him to come out with a class on it.

Last I knew these two videos were not on NPK University but they are on YouTube and helped me a lot:

Full Plant Nutrient Rundown by Harley Smith (Part 1 of 2)
https://www.youtube.com/watch?v=BleWpBYweHE

Harley Smith, Plant Nutrient Research Developer for RAW Nutrients breaks down and talks about each individual plant nutrient and how to utilize them in your garden.
06:44 Nitrogen
12:55 Phosphorus
20:50 Potassium
23:48 Kelp/Seaweed Extract
27:00 Humic Acid
30:30 Fulvic Acid
36:40 Silica
41:10 Yucca Extract
48:25 B-Vitamins

Full Plant Nutrient Rundown by Harley Smith (Part 2 of 2)
https://www.youtube.com/watch?v=zdkwTo79NXE

0:25 Calcium/Magnesium
02:40 Amino Acids
13:20 Cane Molasses
20:30 Questions/Answers
 
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I am very familiar with my compressed gases and their safety protocols, that is why when I saw that I was quoting it exactly... Sorry but as any space buff knows the Apollo 1 astronauts burned up alive on the ground because their capsule was filled with compressed oxygen, and it changed the future of spaceflight forever because of it. Seriously above all else I am #1 when it comes to doing it right and doing it safe, and I know better than to ever use oxygen inside, and if those who need oxygen for medical purposes were smart they would hang the "Oxygen In Use" sign on their front door.

Yes I know my basics of the hormones from cloning and basic biology, but as you so clearly pointed out different ones are needed for different things, and as I continue to read you can clone an entire plant from pretty much any part of the entire plant including its pollen. Which is exactly why I came to this point of wanting to be able to learn to do this, and I realize that this is going to be a lot more researching and reading to learn what I am going to need to know. Thank You for that website, that is exactly the kind of place I was looking to buy my things from where you can get everything you need and it is exact and precise in what you are buying especially when it comes to a medium or something like hormones. I do know enough to know there are different mixes and grades of things and those little tiny differences really can make a serious difference. Now what I really need is to find me a book or detailed website that teaches me the individual and proper steps...

Here is my plan of how I'm planning at going about this entire process... first I have been curious about this subject I've done a few months of researching on it, reading, asking questions, and even found a book to buy on the subject to do even more detailed reading on it., I've also watched the step by step video's by the company who makes the first kit so I got the basics of what I get if I buy their kit. I noted like you saw "medium" they don't say its Agar, they don't specify a specific anything for reference so if there is different grades of blends of it you still won't know what to buy and can't replace anything other than the entire kit, because it contains everything you need. The 2nd kit gives you a bit more details and lets you reorder some things, but again it seems like they are keeping you over the barrel by not telling you everything. Like you pointed out I may need different hormones and stuff for different parts of a plant, a different species of a plant... what are these kits specifically set up to do? Do all the kits grow all plant material? Lots of questions here... I read you can use pollen to Tissue Culture from, does it take a lot of pollen, different hormones? Right now I am at the point I am filled with questions... hence why I have come looking for others who like yourself have done this before, as I am hoping you can tell me websites to get my materials from, perhaps a book to read, a video to watch, a long article I need to go read... I am seriously willing to go put in the work, I am not asking anyone to come hold my hand and hold on while I go do Step #33... and come back now what?

You pointed out I needed a 15-lb pressure cooker, I assume that being it is in the "lab area" it will be being heated by a hot plate to carefully be able to control the heat more perfectly? Oh and that was another question, the bunsen burner, isn't it just going to be being used to sterilize and so my question then comes can I use an alcohol burner instead? I have also been looking more into those DIY laminar airflow cabinets and had some questions if they really are designed like the commercial one's you buy? I mean I am all for DIY and using what you have already, unless it comes at the cost of not doing it right.

At the end of the day before I start buying kits or anything else towards this end goal, I want an entire parts list compiled complete with prices, a step by step set of instructions to follow from A-Z so I can double check my parts list, I want to have at least one good book read on the subject and feel I have a firm grasp of everything needed of me. Then I will make good and sure I feel confident enough to guarantee success, as I will make sure I have resources in place to turn to encase things don't work out so I can figure out what I did wrong and fix it. Then and only then will I start spending the money towards this goal of being able to do it, because otherwise it's a waste of time, money and energy if I can't buy all of this full well knowing I am going to be guaranteed I will reap the rewards in the end for all of this hard work and effort.
 
Oh believe me I am not easily deterred... if I want to know something I sit down ask some people some questions, read at least one book or more on it, watch YouTube videos, scour the web looking for more information on it and then do it... I don't care what the topic or subject is, either I am going to figure it out and be able to do it or I won't... either way at least I will have put the serious and sincere work into trying to do it, which is often more times more work then those who are criticizing me have done.
 
funny story...one of the best friends i ever had was a new-to-me mechanic i took my car to for a simple repair; he was rec by a friend. the insides of my car were all removed because that was a city that routinely flooded in like 45 min so my first month living there my car got flooded while parked. so i had to take out all the interior, seats included, all the carpet padding, all of it down to the metal...then suck out all the water any way possible, then baking soda it to prevent future smells. anyway, he took one look at the interior and was like "what happened here?" i told him. and he was all like, wait, what? because naturally since i'm a girl, i could not have done such a thing....much less be seemingly doing it in exactly the manner in which it should be done in order to be fully successful... we became bffs that day.
 
Yeah I actually had to teach a girl I know outside of school auto mechanics a few years back because the school wouldn't let her into automotive class, because "she was a girl and girls don't take that class". So while she classmates worked on engines, we rebuilt an engine together that was hers and she knew everything in detail about, which only makes it all the funnier to have a sleeper "semi-street legal" truck on the streets. Nobody took her seriously until they heard the pipes when the clutch came out and it took off... never underestimate what someone can do when they are told they can't...
 
Pressure cookers come in 2 different kinds. One has built in heating and are not suitable for tissue culture and the other variety needs placing on a hotplate and their pressure is controlled by a weight preventing the steam escaping, which governs their working pressure. Yes you could use a metho burner. You are interested in doing pollen culture, which is a very advanced area of to be working in. For something that technical you would need access to a university grade research lab and even then, being a novice your chances of success would be almost zero. Why are you interested to pollen culture and what do you hope to achieve? If you look closely at the website I mentioned you will also find that their is also a lot of information about what you need and how to set up a lab.
 
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Yep I have the type in which goes on the stove top/hot plate and then has the weights to set it for 5-10-15 lbs of pressure. My interest in pollen culture is simply the fact that it can be done, that is the problem with where I am at during this stage... I read and read and hear "you can do Tissue Culture with pollen, roots, stem, leaf material etc..." but they don't say anything beyond "you can do it". The problem is you can go to a store right now and buy a transmission rebuild kit for your car and I know plenty of people who have tried only to have to take the mountain of parts into the shop to have someone who knows what they are doing put it back together correctly because the directions only take you so far and don't tell you half of the steps of the things you already need to know. Hence why I am so very glad you are taking the time to help me, because I am only reading "you can do this..." it isn't telling me like you are that I would need a dedicated research lab to accomplish the work at hand... Hence why I am at the drawing board trying to look at everything in detail to get this all figured out on paper before I move forward.
 
We are talking about 2 completely different areas of tissue culture. Basic culture is quite straight forward and simple and most people would be able to achieve reasonable results with practice. Pollen culture on the other hand is very complicated and requires a lot more equipment and knowledge.
The question you need to ask yourself is not what can be done but what you want to do. Unless you have a clear idea on exactly what plants you want to culture and why you are setting yourself up for failure. I am not trying to discourage you but being realistic and offering my oppion. I cannot stress enough the fack that you will need to know exactly what you want to do.
 
I know you and I are on the same point... but I don't think you are seeing that... I don't know enough yet to know that pollen culture and tissue culture are different, or that you need more equipment to do it... All I know is the little that said "you can use Tissue Culture to do...". I am still in the stage of wanting to understand and know what all can I do with this? How practical is it, what equipment is needed to do it, how hard is it to do... I am seriously loaded full of tons of questions about everything from "What is Plant TIssue Culture and What All Can You Do With It?". I don't care if it can be done with pollen, if you need a research lab to do it... I would at best have a small lab of my own where I would grow explants to keep in stasis. What kinds of plants? I don't know, what plants can I grow here for food and medicine that I can't or don't have in abundance, what plants could I sell and turn this knowledge into a cash turn around? Would this help me to get my greenhouse up and going to start my business up so I can one day grow big enough to be a nursery for plants and trees? Thus far I know only that the basics of what I am reading regarding this topic, I don't know a lot about it yet... that is why I keep trying to get you to see and understand that is why I came here to ask questions of where do I start and if anyone else has ever done it? It seems really cool, but I am the first to say "I DO NOT KNOW ENOUGH ABOUT THIS TOPIC AND SUBJECT YET" to be able to do it, know completely what it is, etc...

To try to give you a comparison "It would be like someone saying they know hydroponics means to grow plants in water"... that is a far cry from knowing how to run a hydroponic system to grow plants successfully and know about the nutrients, lighting, etc... I only know the basics of what this is, however if this was the topic of hydroponics I'd just do a search find sites like this one and I'd have people crawling over one another to help me learn everything you need to know about hydroponics. However when it comes to the subject of Tissue Culture it jumps into all kinds of subjects, cell division and not just those relating to plant tissues either, and there is no step by step beginner's guide. I am not even sure if this book I am waiting on is going to put any of this into perspective, I know it sounds cool... I know they sell the DIY Home Kits for it, and I know that it takes equipment and more knowledge than I currently have... stuff I am trying to learn and figure out what I can do with what... so how can I know exactly what I want to do with it, when I don't even know what all I can really do with it because I can't seem to find a site anywhere that outlines it for me. Believe me I keep trying to find a site to outline it, I keep trying new keywords for my search terms, and I keep responding trying to tell you this.
 
To tell you the truth I don,t know what else I can tell you.or how I can help. My only suggestion would be to go to a local university and access their library and talk to their biology department.
 
i don't know if this will help or not but as a former employee of 4 different universities and as a former graduate student i can say w/100% confidence that if you can just find the right person or people who are *SO SO* INTO this they will talk your ear off about it if you want them to. i know that's funny but i don't mean it to be. i promise it's true. so along those lines i'm just gonna throw this out there.... why don't you search for "expert author professor tissue culture cloning" or some combination thereof. alternatively, insert the word conference in there and see what you get. if you can find the tiny little group of academics for whom this topic is pretty much their *entire* life (grad students especially) then you could just write to them directly and ask how to get started, book recs, whatever you like. again, i don't know if you've done this already but just thought i'd put it out there...
 
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