|
Some of these posts are very old and might contain outdated information. You may wish to search for newer posts instead.
|
NecroMyce
Mostly Ghostly



Registered: 05/12/13
Posts: 747
|
Cloning talk
#19348149 - 12/30/13 07:36 PM (10 years, 1 month ago) |
|
|
So i know you can clone with agar by placing some fresh tissue into a sterlized agar plate and then seleting sectors until you get a nice rhizo growth. And then you can halt that growth to save it over time and then take cuts from the fully colonized agar plate and drop it into sterlized rye grain and ive read it takes 8-15 days rather than spore inoculation taking 15-45+ days to get fully colonized, then i usually do grain to grain after that and thatll take another week, thus a whole grow taking 2-3months with rye and monotubs. which is chill but id love to speed it up and maximize my use. i understand agar is a great choice and i plan to take it. But i have a few questions...
Can you take a freshly cut sample from a harvested shroom and drop it into a sterlized rye jar and colonize that? and then do grain to grain? and if so how long would that take? would it be faster than spore inoculation and be considered cloning?
and if anyone has any cloning teks to the like, please post links. i do know there is a search bar and do use it. just like to see what other people do and say currently.
Instead of glass petri dishes, i plan to just use 1/2 pints as agar plates and a baby bottle, or this one bottle with this nice teriyaki sauce in it im almost done with it haha.
|
cronicr



Registered: 08/07/11
Posts: 61,436
Loc: Van Isle
Last seen: 2 years, 8 days
|
|
 the contam rate is extremely high with dropping tissue in grain jars and yes the time factor for the first part is there but once you have a master culture to work with you don't need to go through all the stuff in the beginning like sectoring and isolating and your culture can last years if not more, and not all clones will sector sometimes you get a mono culture right off the hop(best case scenario)
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
|
NecroMyce
Mostly Ghostly



Registered: 05/12/13
Posts: 747
|
Re: Cloning talk [Re: cronicr]
#19348196 - 12/30/13 07:44 PM (10 years, 1 month ago) |
|
|
a mono culture right off the bat?
|
cronicr



Registered: 08/07/11
Posts: 61,436
Loc: Van Isle
Last seen: 2 years, 8 days
|
|
lol yep it happens
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
|
HypnotoadCroaked
Retired, but will check MSGs

Registered: 01/05/13
Posts: 1,168
|
|
Quote:
CaspuuuR said: Instead of glass petri dishes, i plan to just use 1/2 pints as agar plates and a baby bottle, or this one bottle with this nice teriyaki sauce in it im almost done with it haha.
Im going to brush over your statement about dropping tissue samples into grain. Yes it works, but your chance of contamination with even the best kept mushroom are pretty high. In the last 3 months, I am averaging about 50-60% for totally clean clone on agar. The other 40-50% of the time there is contamination. the method you described can work, but you recognize agar is going to be a better asset.
I am a very fond user of "No-pour" petri dishes. I almost exclusively use 1/2 pint wide mouths with plastic lids for this process, and it works great.
Here is a link to a VERY Informative thread concerning alternative petri dishes (i.e. No pour) http://www.shroomery.org/forums/showflat.php/Number/19223885/page/2
Quote:
CaspuuuR said: a mono culture right off the bat?
Most clones USUALLY have a varied genetic makeup. Most have more than a few sectors. IF you isolate a single set of genes on agar, it is called a "Mono culture", and simply is one genetic set, versus the MANY that make up a multi-spore inoculation (MSS).
|
NecroMyce
Mostly Ghostly



Registered: 05/12/13
Posts: 747
|
|
So if i got a specific genetic cloned and cut a piece and dropped it into grain i have a 50-60% chance of success basically?
|
cronicr



Registered: 08/07/11
Posts: 61,436
Loc: Van Isle
Last seen: 2 years, 8 days
|
|
i would say you would be lucky to get that
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
|
tripdawg420
low life with no life



Registered: 02/02/09
Posts: 7,071
Loc: illinois
Last seen: 7 hours, 24 minutes
|
|
Quote:
CaspuuuR said: So if i got a specific genetic cloned and cut a piece and dropped it into grain i have a 50-60% chance of success basically?
like a muchroom tissue or on agar i have never had a piece of mushroom tissue straight to grain work for me ive tryed it prolly 40 times
|
NecroMyce
Mostly Ghostly



Registered: 05/12/13
Posts: 747
|
|
i was talking an agar wedge, whats the possibility of that colonizing when transferred to rye jars.
|
cronicr



Registered: 08/07/11
Posts: 61,436
Loc: Van Isle
Last seen: 2 years, 8 days
|
|
...my bad bud, those chances can be 99.9 if done right
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
|
tripdawg420
low life with no life



Registered: 02/02/09
Posts: 7,071
Loc: illinois
Last seen: 7 hours, 24 minutes
|
Re: Cloning talk [Re: cronicr]
#19349289 - 12/30/13 11:46 PM (10 years, 1 month ago) |
|
|
ya
|
cronicr



Registered: 08/07/11
Posts: 61,436
Loc: Van Isle
Last seen: 2 years, 8 days
|
|
make sure you make a transfer away from the original fruit to make sure it's clean
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
|
NecroMyce
Mostly Ghostly



Registered: 05/12/13
Posts: 747
|
Re: Cloning talk [Re: cronicr]
#19349297 - 12/30/13 11:52 PM (10 years, 1 month ago) |
|
|
Quote:
cronicr said: make sure you make a transfer away from the original fruit to make sure it's clean
What do you mean?
and i guess thats good then.. cuz i was gonna say, then whats the point of agar? haha? but i want to try agar somehow to speed up my colonization process and see some cool agar growth maybe even get a nice even flush.
One thing i do want to ask is. In my grow room i have a shelf. Should i store my colonizing agar plates in there? I personally feel like i shouldnt since i shouldnt keep colonizing spawn in the same room as colonized spawn. Could i put them in my incubator? or maybe a random drawer in my clothes dresser that i can clear out?
|
cronicr



Registered: 08/07/11
Posts: 61,436
Loc: Van Isle
Last seen: 2 years, 8 days
|
|
well when you take a piece of the fruit and put it on agar you let it grow out enough to make a transfer to a new dish, i don't like to use the dish that the fruit was on for grain inoculation
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
|
NecroMyce
Mostly Ghostly



Registered: 05/12/13
Posts: 747
|
Re: Cloning talk [Re: cronicr]
#19349360 - 12/31/13 12:17 AM (10 years, 1 month ago) |
|
|
Basically saying you like to transfer a sector or two first before cutting a wedge and transferring it to a grain jar?
|
cronicr



Registered: 08/07/11
Posts: 61,436
Loc: Van Isle
Last seen: 2 years, 8 days
|
|
exactly
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
|
HypnotoadCroaked
Retired, but will check MSGs

Registered: 01/05/13
Posts: 1,168
|
Re: Cloning talk [Re: cronicr]
#19349386 - 12/31/13 12:26 AM (10 years, 1 month ago) |
|
|
Quote:
CaspuuuR said: So if i got a specific genetic cloned and cut a piece and dropped it into grain i have a 50-60% chance of success basically?
Quote:
cronicr said: well when you take a piece of the fruit and put it on agar you let it grow out enough to make a transfer to a new dish, i don't like to use the dish that the fruit was on for grain inoculation
I don't either. I prefer to make a few xfers. My latest clone had a total of 3 samples taken. One failed bacterial totally. Another got it on the edges, and the 3rd one was "perfect". The partially corrupt culture was xfered in 2x transfers to be safe. Its on grain(s) right now....
Both survivors showed me a boat-ton of genetics. I was basically looking at limited genetics, and nothing close to a monoculture.
The final one was a bit of a goofy thing. It was perfect, with no contamination that I could tell. I wanted to keep all the genes so I cut a "doughnut" around the whole outside of the culture, and relocated sections to 6 jars. 5 out of 6 are fully colonized right now in a mono that is beginning to pin....The other jar made it to the mini mono/straw test bed.
I do not in any way think that what I did was sane, I just felt that I had a good enough handle on it....And TBH, I am still now not 100% comfortable that this will turn out. It was totally arrogant of me to do what I did, but I felt that I got it. I am afraid that the "mushroom gods" will smite it down in its prime for my actions lol. It has proven to be the slowest colonizer I have ever seen. Basically I hate everything about it right now. Its slow and I don't trust it....It is at 100% and fruiting however....Pin porn next week or bust lol.
|
NecroMyce
Mostly Ghostly



Registered: 05/12/13
Posts: 747
|
Re: Cloning talk [Re: cronicr]
#19349400 - 12/31/13 12:28 AM (10 years, 1 month ago) |
|
|
So from my understanding what i can do is. Sterilize my pour jar, and my agar dishes, then PC my agar mix. pour them inside my jars, let them cool. I can then drop wedges freshly harvested from my monotub inside the agar jars with the agar mix inside. I should try my best to be as sterlize as possible while getting a cut from the inside of the stipe to drop inside the agar jar. I let that colonize and try to pick the best 1-3 or however many i want, the best sectors to transfer to another agar jar. and transfer that and either cut a wedge of that and drop it in a rye jar, or cut another sector if it isnt rhizo enough or a monoculture, and then transfer to rye. and it should take 5-15days to get fully colonized inside the jar, rather than taking the time of spore inoculation?
If i was to mix up some agar and made a little to much is there a way to sterily store my agar mix so that i can save it a few days to a week or so to transfer a sector to it? or is it easy enough to just mix some up and PC it the day before i make the transfer?
|
magickspore



Registered: 12/11/12
Posts: 798
Loc: Center of the universe.
Last seen: 8 months, 4 days
|
|
Yes. It replaces MS inoculation to grains. Agar ensures clean cultures. As a benefit you can isolate it down to achieve some pretty amazing pin sets among other great genetics and oddities. If its in something that can be pc'd just re sterilize let cool inoculate.
|
NecroMyce
Mostly Ghostly



Registered: 05/12/13
Posts: 747
|
|
best to clone from MS or best to clone from a fresh fruit?
|
cronicr



Registered: 08/07/11
Posts: 61,436
Loc: Van Isle
Last seen: 2 years, 8 days
|
|
a fresh ms fruit works great
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
|
cronicr



Registered: 08/07/11
Posts: 61,436
Loc: Van Isle
Last seen: 2 years, 8 days
|
Re: Cloning talk [Re: cronicr]
#19351718 - 12/31/13 04:08 PM (10 years, 30 days ago) |
|
|
in fact i can't even think of any other reason to clone anything but a ms fruit
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
|
NecroMyce
Mostly Ghostly



Registered: 05/12/13
Posts: 747
|
Re: Cloning talk [Re: cronicr]
#19351869 - 12/31/13 04:51 PM (10 years, 30 days ago) |
|
|
yeah thats why i asked. because whats the point of using agar if you inoculate with MS lol.
|
magickspore



Registered: 12/11/12
Posts: 798
Loc: Center of the universe.
Last seen: 8 months, 4 days
|
|
As I said it ensures a culture is clean especially useful with wild prints. That's where ms to agar is mostly applied. IMO
|
cronicr



Registered: 08/07/11
Posts: 61,436
Loc: Van Isle
Last seen: 2 years, 8 days
|
|
i go ms to agr transfer a couple times-fruit it out and clone my favorite transfer sectors all over again until it's clean and fruit yet again to se where i'm at and if i'm satisfied put a piece to a slant(which hasn't happened for me for awhile) lol
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
|
NecroMyce
Mostly Ghostly



Registered: 05/12/13
Posts: 747
|
|
Quote:
magickspore said: As I said it ensures a culture is clean especially useful with wild prints. That's where ms to agar is mostly applied. IMO
What do you mean it ensures a clean culture.
|
cronicr



Registered: 08/07/11
Posts: 61,436
Loc: Van Isle
Last seen: 2 years, 8 days
|
|
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
|
Pastywhyte
Say hello to my little friend



Registered: 09/15/12
Posts: 37,810
Loc: Canada
|
|
Spore prints and syringes are rarely clean. Agar is the only guaranteed clean inoculate because you can actually see if its contamed and then transfer away from the contam.
|
magickspore



Registered: 12/11/12
Posts: 798
Loc: Center of the universe.
Last seen: 8 months, 4 days
|
|
|
tripdawg420
low life with no life



Registered: 02/02/09
Posts: 7,071
Loc: illinois
Last seen: 7 hours, 24 minutes
|
|
Quote:
Pastywhyte said: Spore prints and syringes are rarely clean. Agar is the only guaranteed clean inoculate because you can actually see if its contamed and then transfer away from the contam.
|
|