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OfflineTh3Issu3
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Registered: 12/30/14
Posts: 671
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Last seen: 2 months, 24 days
No Issue Easy Agar TEK * 1
    #23967419 - 12/30/16 07:30 AM (7 years, 2 months ago)

No Issue Easy agar

In this section I’m going to go over how I make agar plates. I will start with the materials that you will need and where to purchase them. Then explain my sterile technique and pour. This will be first in my journal of No Issues  Cultivating.
* * Now most of these,  if not all of them,  you can find at Walmart or any number of local stores. If you are like me and don’t want a thousand questions on what you are doing from the meth head cashier, you will probably opt to pick a few things up online.  Almost everything that I have on this list you can search on Amazon.com and find using the key works that I used.**


                            Materials
1. Potato Dextrose Agar (PDA)
2.  20 Pack Sterile Plastic Petri Dishes, 100mm Dia x 15mm Deep, with Lid
3. Pint jars with lids, (2)
4. Plastic Saran wrap
5. Tin foil
6. Scissors
7. SAB / Flow Hood
8. Pressure Cooker
9. Small Cooking pot
10. Swan 70% Isopropyl Alcohol
11. Disposable Nitrile Exam Powder Free Gloves
12. Dawn Dish soap
13. Paper towel
14. Digital Scale
15. Towel



                    Agar prep. And sterilization

First thing you will notice is that the directions are stated on the bottle.  These are followed in the steps below.



1. Take your pint jar and use it to measure the correct amount of water. I fill it up to the 200ml mark twice then the 100 ml mark.  ALWAYS USE COLD WATER!
2. Now pour the water into your small pot on the stove.
3. Take your scale out, flip the Agar bottle cap upside down on the scale and pour some flakes into it. I usually do this at 5 grams a time to equal 20 in total. Then pour it into your pot of 500ml of cold water.
4. Turn the heat on med-high and stir the contents to evenly mix with the water.
5. Continue to stir until your solution reaches a boil for one minute.
6. Cut your heat off.
7. Pour your Potato Dextrose Agar solution into two pint jars. It should read about 250 ml in each.
8. Now put the lid on and tighten the top almost all the way shut. Leave it a little loose for removal after The Pressure Cooking cycle.
                                             
          :hairmetal: Congrats. You successfully made Agar Solution! 

                                             


        Now it’s time to move on to the sterilization process.

1. Don’t be a dirt bag, wash and dry your jars with dish soap first. Always!
2. You will need to cut two 4 inch by 4 inch square pieces of tin foil. These go over the top of your jar, so as to not let any water splash up into your jar and solution while your pressure cooker is sterilizing your new made agar solution.
3.  Now fill up your Pressure cooker to the factory recommended fill point using room temperature tap water.
4.  Put your jars in the Pressure Cooker in the center with a small space in between them.
5. Now place your cover on the pressure cooker without the weighted rocker.
6. Change your stove temp. to the setting “7” or med high.
6. Let the steam come from the pressure cooker for a good 5-10 minutes.
7. Place the 15 psi rocker back on the port and wait until it starts to shoots steam out while rocking back and forth.
8. Now set the timer for 45 minutes.
9. After 45 minutes turn off your stove.
10. Leave the rocker on and let depressurize for about 2-3 hours.
11. Remove the Pressure Cooker and bring it to the area you will be working in. Keep the lid and rocker on and set it on a towel. Your pressure cooking will still be warm.
                               
                                                                 


                              Agar Pour Prep

Depending on what you use for example, a Flow Hood or a SAB. Skip to the corresponding section.


                            Flow Hood Prep.
1.  Turn on your flow hood and let run for 10-20 minutes as to let is scrub the air in your room. Ensure the door in your room is closet.
2. Put on your gloves and clean your hand with the isopropyl alcohol.
3. Cut your clear plastic saran wrap in strips measuring about 1 inch by 6 inches. Then put to the side for later use.
4. Squirt the Isopropyl alcohol on a paper towel and scrub the table your Flow Hood sits on as well as everything in the air stream.  Be generous with the solution.
5. Clean your scissors with the Isopropyl alcohol. Make sure to clean all of them, not just the blade.  Then place in your flow hoods air stream.
6. Wipe down the packaging of your petri dishes and cut the bottom of the package.  Place them in front of your flow hood ensuring the bottom of the dish or the receiving part of the dish is sitting down.
6. Remove the Pressure cooker lid. Take the two jars of Potato Dextrose Agar solution give them a little shake in a circular motion to mix the settled solution and place them in front of the flow hoods airstream.





            SAB PREP.


1.  Ensure there are no fans on, and the windows are closed in the room you are working in. You want the air to be as still as possible in your work area.
2. Put on your gloves and clean your hands with Iso alcohol.
3. Place your SAB on a clean towel, sprayed with soapy water. 
4. Completely wipe the area inside as well as out with the iso alcohol on a paper towel. Then mist with the solution inside the SAB spraying twords the top and letting the mist fall down.
5. Cut the plastic Saran Wrap into 1 inch by 6 inch strips and place inside your SAB.
6. Now put the scissors, petri dishes, and alcohol with a few paper towels in your SAB.
6. Remove the Pressure cooker lid. Take the two jars of Potato Dextrose Agar solution  give them a little shake in a circular motion to mix the settled solution and place them inside your SAB.
7. Clean your scissors with the Isopropyl alcohol. Make sure to clean all of them, not just the blade. 
8. Wipe down the packaging of your petri dishes and cut the bottom of the package.  Place them in front of your flow hood ensuring the bottom of the dish or the receiving part of the dish is sitting down.



                                                                                                        AGAR POUR

              **Now regardless if you are using a SAB or a Flow hood the pour will be the same. **

1.  Wash your gloves off with the alcohol and arms up to where your clothing starts.
2. Scrub the jars you are using with an alcohol soaked paper towel.  Take your time with this and make sure you scrub every square inch.
3. Now remove the metal rings from the jars and set aside. Scrub the area where the ring was with a paper towel and alcohol solution.
4. Now grab the top of the bag that the petri dishes are in and lift up to expose 3 to 4 dishes.  Then place the bag holding the other petri dishes next to the ones that are out.
5. (If you are right handed ) Place your left palm over the 3 – 4 petri dishes that are out of the bag and grab the top lid of the bottom dish.
6. Remove the jar lid.
7. Pick up the lid of the receiving dish and pour the warm agar solution into the dish, filling it half way. 
8. Still holding onto the others, place the lid back.
9. With the next dish you do the same thing but bottom rest on the first dish and so on. I usually only do about 3 - 4 at a time. These all sit on top of each other in a nice neat stack.
10. Just repeat the process on steps 4 – 9 until all the solution is gone.
11. After this is complete you will take the Plastic saran wrap strips and put the around your petri dish to form a seal.

                                                                                            :hairmetal:You’re done!:hairmetal:
** What I do is wait about a week before I use any dishes, so I can ensure that they were poured clean before I use them ** .

Edited by Th3Issu3 (12/30/16 09:31 AM)

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OfflineTh3Issu3
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Registered: 12/30/14
Posts: 671
Loc: fucktown
Last seen: 2 months, 24 days
Re: No Issue Easy Agar TEK [Re: Th3Issu3]
    #23967421 - 12/30/16 07:32 AM (7 years, 2 months ago)

This is my first TEK and admission to my journal. This new year I plan on giving back to the community what you guys gave me and hopefully help a few people along the way.

I appreciate any feedback! 

Thanks.

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Offlinetump
ban the undead
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Registered: 03/17/16
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Re: No Issue Easy Agar TEK [Re: Th3Issu3]
    #23967431 - 12/30/16 07:43 AM (7 years, 2 months ago)

Neato

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Offlinegermanauslander
Forever Learner

Registered: 11/06/16
Posts: 100
Last seen: 5 years, 7 months
Re: No Issue Easy Agar TEK [Re: tump]
    #23967497 - 12/30/16 08:40 AM (7 years, 2 months ago)

Good write up. If someone wanted to follow this but use malt extract in lieu of the PDA would that work?  With the appropriate proportions for Malt Extract Agar.

Also what if all that a person has available is quart mason jars, would that work instead of the pint jars.

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InvisibleSubnet Mask
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Re: No Issue Easy Agar TEK [Re: germanauslander]
    #23967505 - 12/30/16 08:44 AM (7 years, 2 months ago)

I want issues in my agar mix so this isn't going to work for me.

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OfflineAK1000
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Re: No Issue Easy Agar TEK [Re: Th3Issu3]
    #23967567 - 12/30/16 09:17 AM (7 years, 2 months ago)

Quote:

Th3Issu3 said:
            SAB PREP.


1.  Ensure there are no fans on, and the windows are closed in the room you are working in. You want the air to be as still as possible in your work area.
2. Put on your gloves and clean your hands with Isopropyl alcohol.
3. Place your SAB on a clean dry towel.
4. Completely wipe the area inside as well as out with the isopropyl alcohol on a paper towel.




Your SAB prep seems to be lacking. The SAB sits on a towel, yes, but not dry. I have a spray bottle filled with soapy water which I just pour the whole thing out usually onto the towel until every inch is soaked.  Then I refill the bottle with some soap and water and spray the inside walls until every inch of the inside walls in every direction are wet. No need to use alcohol on the inside walls, but you DO want to have everything wet.  I also spray inside it every few minutes.  Also, you want to spray from the top so that the mist falls down and brings other particulates in the air down with it.


--------------------
-AK

I'm a noob and :heart: this forum. Here's What I've Learned My First FOUR Months.

Best resources: Bods Easy AF List / Ultimate Tek Compendium

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OfflineTh3Issu3
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Registered: 12/30/14
Posts: 671
Loc: fucktown
Last seen: 2 months, 24 days
Re: No Issue Easy Agar TEK [Re: AK1000]
    #23967572 - 12/30/16 09:18 AM (7 years, 2 months ago)

Quote:

AK1000 said:
Quote:

Th3Issu3 said:
            SAB PREP.


1.  Ensure there are no fans on, and the windows are closed in the room you are working in. You want the air to be as still as possible in your work area.
2. Put on your gloves and clean your hands with Isopropyl alcohol.
3. Place your SAB on a clean dry towel.
4. Completely wipe the area inside as well as out with the isopropyl alcohol on a paper towel.




Your SAB prep seems to be lacking. The SAB sits on a towel, yes, but not dry. I have a spray bottle filled with soapy water which I just pour the whole thing out usually onto the towel until every inch is soaked.  Then I refill the bottle with some soap and water and spray the inside walls until every inch of the inside walls in every direction are wet. No need to use alcohol on the inside walls, but you DO want to have everything wet.  I also spray inside it every few minutes.  Also, you want to spray from the top so that the mist falls down and brings other particulates in the air down with it.




Thank you...Im adding that in now!

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InvisibleMycolorado
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Re: No Issue Easy Agar TEK [Re: AK1000]
    #23967592 - 12/30/16 09:25 AM (7 years, 2 months ago)

Quote:



Your SAB prep seems to be lacking. The SAB sits on a towel, yes, but not dry. I have a spray bottle filled with soapy water which I just pour the whole thing out usually onto the towel until every inch is soaked.  Then I refill the bottle with some soap and water and spray the inside walls until every inch of the inside walls in every direction are wet. No need to use alcohol on the inside walls, but you DO want to have everything wet.  I also spray inside it every few minutes.  Also, you want to spray from the top so that the mist falls down and brings other particulates in the air down with it.




Seems a bit excessive.  I know we all work in different climates and environments with varying levels of contaminants, but this just seems like a soppy mess to clean up later.  I'll give a couple spritz of alcohol in the SAB but that's it.

Edited by Mycolorado (12/30/16 09:28 AM)

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OfflineTh3Issu3
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Registered: 12/30/14
Posts: 671
Loc: fucktown
Last seen: 2 months, 24 days
Re: No Issue Easy Agar TEK [Re: Mycolorado]
    #23967599 - 12/30/16 09:27 AM (7 years, 2 months ago)

I kind of simplified it a little.

1.  Ensure there are no fans on, and the windows are closed in the room you are working in. You want the air to be as still as possible in your work area.
2. Put on your gloves and clean your hands with Iso alcohol.
3. Place your SAB on a clean towel, sprayed with soapy water.
4. Completely wipe the area inside as well as out with the iso alcohol on a paper towel. Then mist with the solution inside the SAB spraying twords the top and letting the mist fall down.

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OfflineTh3Issu3
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Registered: 12/30/14
Posts: 671
Loc: fucktown
Last seen: 2 months, 24 days
Re: No Issue Easy Agar TEK [Re: germanauslander]
    #23967615 - 12/30/16 09:34 AM (7 years, 2 months ago)

Quote:

germanauslander said:
Good write up. If someone wanted to follow this but use malt extract in lieu of the PDA would that work?  With the appropriate proportions for Malt Extract Agar.

Also what if all that a person has available is quart mason jars, would that work instead of the pint jars.





To be honest I've never used malt extract agar and completely unfamiliar with it. I'd assume if it is anything like PDA you'd add the recommended amount per ml of water then follow the steps as stated above to prepare the solution, but like i said im not familiar with using it.

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OfflineMoonlightblue
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Re: No Issue Easy Agar TEK [Re: Th3Issu3]
    #23967641 - 12/30/16 09:47 AM (7 years, 2 months ago)

I've used PDA once, and had really slow growth. The potato makes the agar too solid. And it's hard to see mycelium bcus potato is white-ish too.

Malt extract is the best in my opinion. It's makes clear like solution, easy to see the myc and faster growth. IME

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OfflineMoonlightblue
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Re: No Issue Easy Agar TEK [Re: germanauslander]
    #23967649 - 12/30/16 09:52 AM (7 years, 2 months ago)

Quote:

germanauslander said:
Good write up. If someone wanted to follow this but use malt extract in lieu of the PDA would that work?  With the appropriate proportions for Malt Extract Agar.

Also what if all that a person has available is quart mason jars, would that work instead of the pint jars.




150ml tap water
3g agar
3g mea
1g dextrose (optional)

Yes, mason jars is all I use as petri dishes with great success.

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OfflineAK1000
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Re: No Issue Easy Agar TEK [Re: Mycolorado]
    #23967777 - 12/30/16 10:43 AM (7 years, 2 months ago)

Quote:

Mycolorado said:
Quote:



Your SAB prep seems to be lacking. The SAB sits on a towel, yes, but not dry. I have a spray bottle filled with soapy water which I just pour the whole thing out usually onto the towel until every inch is soaked.  Then I refill the bottle with some soap and water and spray the inside walls until every inch of the inside walls in every direction are wet. No need to use alcohol on the inside walls, but you DO want to have everything wet.  I also spray inside it every few minutes.  Also, you want to spray from the top so that the mist falls down and brings other particulates in the air down with it.




Seems a bit excessive.  I know we all work in different climates and environments with varying levels of contaminants, but this just seems like a soppy mess to clean up later.  I'll give a couple spritz of alcohol in the SAB but that's it.




What does prepping a SAB have anything to do with climate? My understanding of the concept of the SAB is to utilize the concept of Still Air. You're not trying to sterilize everything but rather spray everything down inside so there's less shit floating around and just do everything with steady hands and never put your hands over open containers.

I've always been told you need to spray all the walls and everything and make sure it's wet though, and that takes like, what... 15-20 seconds? Hardly excessive. In regards to the soapy mess... my towel and big ass tote with 6"+ holes in it is pretty much only used for a SAB, so I don't need to clean anything up, it just dries up. :shrugs: What may be excessive is my use of soapy water rather than just plain water. Hopefully someone else more experienced than the both of us will chime in.

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InvisibleMycolorado
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Re: No Issue Easy Agar TEK [Re: AK1000]
    #23967822 - 12/30/16 10:56 AM (7 years, 2 months ago)

I just mean there are more airborne contaminants in some climates than there are in others...tropical locale v desert.  I'm not trying to talk shit to you, just stating that your method seems a bit excessive is all.  I live in an arid environment and don't get many contams...If I do, 99.99% of the time it's bacterial and it comes from me or the spores or the syringe.  You can look at some of my threads to see I don't have issues with agar cultures or a lack of experience.  Again, not talking shit, and not trying to jack this thread, just adding my opinion.

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OfflineNDStepp84
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Re: No Issue Easy Agar TEK [Re: Mycolorado]
    #23967922 - 12/30/16 11:34 AM (7 years, 2 months ago)

I wouldn't recommend spraying a chamber with alcohol for a couple reasons, first being you're turning your SAB into a combustion chamber with a burning torch sitting next to it. People here have went to the ER with severe burns doing so. Next the purpose of spraying the walls is so stuff floating in the air sticks to the walls and alcohol evaporates too fast. Soapy water will keep the walls wet way longer.


--------------------

"I am free, no matter what rules surround me. If I find them tolerable, I tolerate them; if I find them too obnoxious, I break them. I am free because I know that I alone am morally responsible for everything I do.
-Robert A. Heinlein 

:takingnotes: Links and teks:takingnotes:
ND's grow log and discussion
Plant thread

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InvisibleMycolorado
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Re: No Issue Easy Agar TEK [Re: NDStepp84]
    #23967999 - 12/30/16 12:13 PM (7 years, 2 months ago)

Yeah, I've got a bottle of alcohol to clean the bench top with so that's what I use...when I say spritz, I mean a quick small spray...I use a butane torch turned down to flame my tools INSIDE the SAB, so there is very little alcohol floating around in there.

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