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#21 Microbe

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Posted 07 November 2016 - 05:00 PM

Its been awhile since i shared my filter design so i figure might as well do it again.

I swear by these filters and they are very cheap to make.

I get micropore tape at CVS at the pharmacy, they keep it behind the counter and not in resale packaging but they hook me up with 6 rolls 2" wide for $4.00. This will last me several years.

I purchase the eazyfelt at hobby lobby. For a 12x14 sheet it costs me $ 1.07 after tax. There are different colors also but i prefer white so i can identify contamination growing on the filter, it has happened.

Eazyfelt is matted polyfill and when used in conjunction with micropore tape, i would put these up against SFD's.

I still use the 5 hole pattern as shown

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A take a piece of micropore tape and lay the felt as centred as possible.

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Place it over the holes and take my fingernail and push the tape down around the perimeter of the felt to make sure there is no gap between the felt and lid. In the PC the filters get wet but then the tape dries like freaking concrete firmly holding the felt in place.

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Fucking brilliant. I like the five hole design. When I run micropore through the PC I have different results though. You use rings under there?
Yes. I used to not but im getting back unto sugar LC's so i purchased some silicone gaskets from Amazon but figured i might as well use them.

That is paint on my hand. I was painting my ceiling that day. Spent my vacation working on a room remodel and it turned out excellent. I wont post it through because my wife already posted it all over Facebook and in that slight chance 1 of her 1,000 friends is here at topia, well my anonymity would jeopardized.

Edited by Microbe77, 07 November 2016 - 05:11 PM.

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#22 CatsAndBats

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Posted 07 November 2016 - 06:29 PM

The gaskets fit right in purrfectly. Great seal on them. Thanks for these posts, real good stuff.


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#23 coorsmikey

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Posted 12 November 2016 - 03:08 PM

@HrVanker What are your thoughts on using colchicine as a chemical method on crossbreeding? Much cheaper and more readily available then rattlesnake venom.


Where did you come up with the idea of using colchicine?
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#24 Microbe

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Posted 12 November 2016 - 05:07 PM

@HrVanker What are your thoughts on using colchicine as a chemical method on crossbreeding? Much cheaper and more readily available then rattlesnake venom.

Where did you come up with the idea of using colchicine?
It is commonly used when breeding plants. Colchicine is known to induce polyploidy a condition in which a normally diploid cell or organism acquires one or more additional sets of chromosomes. In other words, the polyploid cell or organism has three or more times the haploid chromosome number . Polyploidy arises as the result of total nondisjunction of chromosomes during mitosis or meiosis.

What i think it will do is double the haploids in the monokaryotic cultures increasing the probability of comparability allowing them to mate. This is my thought process around it. I could be way off. I also think will help stabilize the newly acquired hybrid.


This may result in inbreeding of a monokaryotic culture but if it did, would it fruit?

If it would then maybe this will be the best way to obtain a monoculture. Monospore germination isolate and treat with colchicine and double the haploids and maybe it will mate with itself!


Anyway im probably going with the monospore germination by streaking so im not sure i will experiment with this or not. It probably makes more sense trying to use it and get a single monokaryotic strain to breed with itself now that i think about it.


Here are a few reads, i skimmed through these but also google double haploid production and hybridization.

http://scialert.net/....2007.2334.2340

https://www.google.c...dGk8HVxrBXALpQQ

http://www.technolog...1210017019.html

http://rspb.royalsoc.../rspb.2012.0434

Edited by Microbe77, 12 November 2016 - 05:08 PM.


#25 coorsmikey

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Posted 12 November 2016 - 05:46 PM

@HrVanker What are your thoughts on using colchicine as a chemical method on crossbreeding? Much cheaper and more readily available then rattlesnake venom.

Where did you come up with the idea of using colchicine?
It is commonly used when breeding plants. Colchicine is known to induce polyploidy a condition in which a normally diploid cell or organism acquires one or more additional sets of chromosomes. In other words, the polyploid cell or organism has three or more times the haploid chromosome number . Polyploidy arises as the result of total nondisjunction of chromosomes during mitosis or meiosis.
What i think it will do is double the haploids in the monokaryotic cultures increasing the probability of comparability allowing them to mate. This is my thought process around it. I could be way off. I also think will help stabilize the newly acquired hybrid.
This may result in inbreeding of a monokaryotic culture but if it did, would it fruit?
If it would then maybe this will be the best way to obtain a monoculture. Monospore germination isolate and treat with colchicine and double the haploids and maybe it will mate with itself!
Anyway im probably going with the monospore germination by streaking so im not sure i will experiment with this or not. It probably makes more sense trying to use it and get a single monokaryotic strain to breed with itself now that i think about it.
Here are a few reads, i skimmed through these but also google double haploid production and hybridization. http://scialert.net/....2007.2334.2340https://www.google.c...dGk8HVxrBXALpQQhttp://www.technolog...1210017019.htmlhttp://rspb.royalsoc.../rspb.2012.0434
Thank you 77! This is something that I have been playing with so to hear you bring it helps back up what I have been thinking. I will take a look at the link provided. And of course I'll share anything I come up too.

Edited by coorsmikey, 12 November 2016 - 05:47 PM.

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#26 Microbe

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Posted 12 November 2016 - 06:09 PM

@HrVanker What are your thoughts on using colchicine as a chemical method on crossbreeding? Much cheaper and more readily available then rattlesnake venom.

Where did you come up with the idea of using colchicine?
It is commonly used when breeding plants. Colchicine is known to induce polyploidy a condition in which a normally diploid cell or organism acquires one or more additional sets of chromosomes. In other words, the polyploid cell or organism has three or more times the haploid chromosome number . Polyploidy arises as the result of total nondisjunction of chromosomes during mitosis or meiosis.
What i think it will do is double the haploids in the monokaryotic cultures increasing the probability of comparability allowing them to mate. This is my thought process around it. I could be way off. I also think will help stabilize the newly acquired hybrid.
This may result in inbreeding of a monokaryotic culture but if it did, would it fruit?
If it would then maybe this will be the best way to obtain a monoculture. Monospore germination isolate and treat with colchicine and double the haploids and maybe it will mate with itself!
Anyway im probably going with the monospore germination by streaking so im not sure i will experiment with this or not. It probably makes more sense trying to use it and get a single monokaryotic strain to breed with itself now that i think about it.
Here are a few reads, i skimmed through these but also google double haploid production and hybridization. http://scialert.net/....2007.2334.2340https://www.google.c...dGk8HVxrBXALpQQhttp://www.technolog...1210017019.htmlhttp://rspb.royalsoc.../rspb.2012.0434
Thank you 77! This is something that I have been playing with so to hear you bring it helps back up what I have been thinking. I will take a look at the link provided. And of course I'll share anything I come up too.
Read this one also, it talks about breeding a single monokaryotic without the need for a mate. It doesnt specify how or the use of it but it would only be possible through double haploid production.

http://www.americorp...-mushrooms.html

Good luck and please share your findings or any research that will be beneficial. Perhaps you can join in on the thread when started!

Edited by Microbe77, 12 November 2016 - 06:10 PM.


#27 Microbe

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Posted 12 November 2016 - 08:27 PM

@coorsmikey

Here is my favorite one, i couldn't find it earlier on my lunch but here it is! NCBI is my favorite fungi science abstract resource followed by Penn State University.

https://www.ncbi.nlm...les/PMC3385130/

Edited by Microbe77, 12 November 2016 - 08:28 PM.

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#28 Microbe

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Posted 13 November 2016 - 02:50 AM

Man o man im so used to complete colonization of single qt jar in 5 days so this is slow. I am incubating at room temp in a cabinet attached to a exterior wall. I typically shoot for target temp of 82° when i have a controlled incubating room or area. Cool temps and weak cultures, its just not as fast as i like. Im ok with it as long as they fruit prolific!

Im not sure im digging that inoculate loop alternative. It is diffidently a awesome tool for cleaning up culture, isolating a mono, and now, capturing a monokaryotic before it gets to bump uglies!

This is one of those boring posts i was talking about my intro. I promise once i get back into the groove, im going to attempt to apply everything i have read about during my absence.

If something needs its own thread, i will start it. This is just a random generic thread so i dont clutter up the forum. I wont bury any teks or innovative breakthroughs in amateur mycology here ;)

Some growth and clean plates.
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No growth and clean.

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Just some slow ass jars, all are clean. I do want to note that i tossed my APE jar because it had some nice growth and about 25% so i shook it, and then it fully colonized in 48 hours! Lmao>mold

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Edited by Microbe77, 13 November 2016 - 04:05 AM.

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#29 peacefrog

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Posted 13 November 2016 - 07:02 AM

Good stuff as always, microbe.
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#30 Microbe

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Posted 15 November 2016 - 12:49 AM

I think im going to have to set up a incubator. I dont like slow growth at all and i was used to working in a closet with a heater plugged into a temperature controller and until i get my electrical upgraded i cant run a space heater the good thing is i did get some breaker style fuses so i guess i could attempt it without worrying having to replace the fuses. Did even know they existed until several months ago!

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3 plates are are still not showing life and they are clean so i will attempt to inoculate again. I may have not touched a agar plate after flaming sterilizing destroying the little bit of mycelium i attempted to transfer. Jars are moving slowly and this is about the point i would shake them and expect full colonization in 2 days before giving the final shake and recovery but thats not happening. I will let them go another 3 days. Nothing special just some wbs getting chewed on my some mycelium.

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#31 CatsAndBats

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Posted 15 November 2016 - 08:10 AM

Are you inoculating with an entire plate per quart jar?



#32 Microbe

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Posted 17 November 2016 - 08:43 PM

Are you inoculating with an entire plate per quart jar?

1/2 plate for most and a few with a whole plate but remember i work with with 60 or 65 mm plates, i dont remember which but they are small.

These cultures are weak also, im probably going to go back to spore and start over. Now if they fruit nicely i will keep them.

I have noticed that WBS jars have always taken a little linger also to colonize then oats because there is a significant more amount of surface area per volume.

Im also used to inoculating with glc which IME gave me fast results comparable to that of a slightly higher them standard ratio of g2g.

Spawn run through jars and subs were always done the low 80's.

All this combined is just presenting to me something im not used to seeing but im in no hurry at the moment.
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#33 Microbe

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Posted 18 November 2016 - 04:23 PM

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Didnt look at the plates today but obviously i checked on the jars. They are beyond shaking and i will just let them finish before shaking the final time. By looking at these cultures i am still disappointed and they are motivating me to go back to spore and start over. These will be ready to be milked soon.

#34 CatsAndBats

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Posted 18 November 2016 - 07:13 PM

Good, if you stop throwing out cultures I'll get plates for the breeding thread.


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#35 whitethumb

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Posted 19 November 2016 - 12:01 AM

looking good microbe
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#36 Microbe

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Posted 19 November 2016 - 01:41 AM

Decided to not be lazy and remove the jars from the top of my culture storage box so i could pull it out and check on them.

To my surprise the 3 plates that had no growth finally do. All plates were clean and i think this single strand nichrome wire is a excellent tool.

I think what helped all of thee above was the upper 70's temps we had here the last 2 days.

All plates are a success and the plates laying across the hinged point of lid are the 3 that i posted twice already showing no growth at all.

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Edited by Microbe77, 19 November 2016 - 01:43 AM.


#37 Microbe

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Posted 02 December 2016 - 01:30 PM

Glc and G2G unknown masters have been done for a week and have recovered a few times. Shook the jars again yesterday in preparation for extracting the mycelium tonight.

Have a bag of oats ready to go and look excellent and will probably start a thread for strain identification of these unknowns as it wont be so random, this is for random shit, but i tend to be random.

Finally im ready to grow!
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#38 Microbe

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Posted 03 December 2016 - 05:36 PM

This is some of the best oats i have ever made. Im always trying to fund ways to improve and i will explain it detail when i do a updated oat prep tek.


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It took about 5 days for these to be inflated enough to work with and those of you who use bags know the pita they can be trying to inoculate and disperse inoculate while still under a slught vacuum but i found a simple way to remedy that. I just pull the bags apart at the fikter and within 24 hours they are food to go.

This sucks working with bags like this!

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Out of 50 bags there were a few runs where i didnt pack my towels right and melted 4 bags so i tossed them in the freezer and have some grain already hydrated to make some glc, of coarse i will sterilize it again. I posted a few years ago about storing hydrated grains in the freezer and was a success. This freezer is dirty, its the garage fridge and we do not take care of it very well as you can see.

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My GLC jars already recovering from the shake the night before, i always shake it the day before instead at the time i need it.

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GLC ready to be extracted.

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I do not typically use WBS and i had a grain clog up ine of my tubes so it ended up being g2g along with the water that was in it. The water is much darker then when i use oats also. Anyway there was some nice biomass in the syringes.

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The last little bit of glc, this is what i call my buddy syringe @tendrfoot!

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The freaking look right for the micropore tape and look back left oh shit where is hole bag! Lmao

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Im g2g about 20 bags also. My process is a little off right now and i am just practicing. Eventually i will get back to agar>glc masters>secondary masters> g2g working spawn.

The freaking outlet being installed is very useful!

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MAD SCIENTIST

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#39 Needles

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Posted 03 December 2016 - 09:55 PM

Your oat bags look awesome Microbe, you say you're garage fridge has a dirty freezer but I would still go for one of those orange freezie pops.... Nice work on the grain masters and the liquid inoculate syringes.....
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#40 Dilatedpupils

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Posted 07 December 2016 - 11:15 PM

This is going to be a bunch of random stuff i will be working on. Some very boring, some very interesting.

Lets start with a happy Halloween!

This picture was taken 2 years ago at my old house, damn i remember working in that hallway right beside my washer and dryer and behind me through that door is where the magic happened!

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About to do some flow bench work and while im doing that i decided to give my pp5 squeeze bottle agar container another go with several modifications. The PC is heating up right now with a test bottle in it with just water.

Remember when i tried this the first time the syringe filters popped off, the bottles floated around and melted? If not, they did!

So a went with a eazyfelt/micropore filter, and also used some tape around the lid, i dont trust it and even though this is just a test run, i want to mimic exactly how it be if i were cooking some agar.

I dropped the bottle in a wide mouth jar to keep it put once the water starts rolling, i cant believe i didnt even think about that my first run!

Sicks though, i had to bring out one of my 24's lmao

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Time to get to work!

Miccccroobbe in dah houuuseeee :) I'm back on brodur:)

life is good:)
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