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Hey all! I on the 25th i inoculated some agar dishes with a summer fruiting mushroom. After 8 days i have seen no growth (mycelial or other) on the plates. The spore prints were dated 2019 and i followed pastywires tek. They are being incubated at ~24oc Any knowledge and wisdom would be greatly appreciated!
Just thought I'd let you know: I have had a 100% sterile success rate with the peroxide agar kitchen tek, both with the cooked method ( no autoclaving ) and the autoclaved method. Methods and comparison below Dispensing and media transfer was done under absolutely not sterile conditions, in order to see if the tek could be replicated anywhere ( those of you who know my kitchen can stop laughing ). I was a tad sloppy with my transfer tek as well to see what would happen Reporting it here because sometimes the signal/noise ratio can be a little loud elsewhere Batch 1- autoclaved + 6ml/L 3% H2O2 Chinese sauce containers were autoclaved in a bag for 20 min 100ml liquid MEA autoclaved for 20 min, consisting of the following 20g/L light malt extract 0.1g/L garden lime 0.1g/L potassium phosphate dibasic 15g/L gelcarin ( 20g/L agar works just as well ) Media was not subject to a pH test Post autoclave the media was allowed to cool and just above setting point 6ml/L 3% H2O2 was added. Media was swirled heavily so that the inner surfaces of the bottle were fully coated Dispensed immediately into sauce containers *on the kitchen bench* in the open air Lids were placed lightly over the containers and 1hr later the plates were completely sealed after inoculation with various species Batch 2- cooked 1hr + 6ml/L 3% H2O2 Chinese sauce containers were autoclaved in a bag for 20 min 100ml liquid MEA cooked for 1hr by placing media container in an open saucepan. Media container lids were left loose. Water came up the the media level- bottles weren't more than 3/4 immersed. Cooked at a slow boil for 1hr 20g/L light malt extract 0.1g/L garden lime 0.1g/L potassium phosphate dibasic 15g/L gelcarin ( 20g/L agar works just as well ) Media was not subject to a pH test Post autoclave the media was allowed to cool and just above setting point 6ml/L 3% H2O2 was added. Media was swirled heavily so that the inner surfaces of the bottle were fully coated Dispensed immediately into sauce containers *on the kitchen bench* in the open air Lids were placed lightly over the containers and 1hr later the plates were completely sealed after inoculation with various species Sterile ( non-peroxide MEA ) library cultures were opened and haphazardly used ( left open for much of the inoculation process ) to inoculate the plates above using a scalpel blade which was only cleaned and flamed between species The following species were placed in the centre of the agar of each container Reishi ( Ganoderma lucida ) Blue Oyster ( Pleurotus spp ) Lion's mane ( Hericium erinaceus ) Elm Oyster ( Hypsizgus ulmanarius ) Plates were sealed with Austraseal and incubated in the dark at 22C 2 plates from the cooked group and 2 plates from the autoclaved group were left uninoculated as controls to check for contamination during dispensing By my judgement it was all a bit haphazard and I didn't believe it would work. Even a contam rate of 10% per batch would have been acceptable At +1 week there is no contamination, anywhere, and growth is good for the Reishi and Elm Oyster. Still waiting on the Blue Oyster and Lion's Mane, but plenty of time yet- those parent cultures could have been a little old- I have storage/ library cultures and can reinoculate from them easily at +3 weeks If you are thinking about the peroxide tek for agar- give it a go. I've only made it sound complex cos I wanted to write it all up so you know I took all the steps. I now pronounce this part of the tek piss easy Edited very fast, because I am an idiot and forgot to put the decimal point in
Darklight posted a topic in MycologyLike it says in the title... Seeking clean slant or plate of Trametes versicolor that has been confirmed by fruiting and is ready to send. Will pay. Dammit I knew I shoulda got one isolated from here back when it was raining enough to make mushrooms :D
Hi, I've recently become a registered sole trader and in support of the community who helped supply me with much needed cultures, I'd like to return the favor. This offer is aimed at amateur mycologists or those interested in mycology without higher end equipment. The Offer: I'd like to offer a cloning (or isolation) service to anyone looking to put a species onto agar. I'll get this out of the way first, this offer is applicable only for species legal to possess live. If you have spores or a wild specimen that you'd like to get onto agar, or need a clone or isolation of an existing culture, PM me or contact me via email at [email protected]. What I ask for the service is that shipping costs be covered by the purchaser and $12.00 to cover the cost of materials for the clone. I remember exactly what it was like to have to make do for equipment, and while it worked well in some situations, there's often something that just doesn't go correctly. While mycology was hobby for me, I was confined to a small glove box working with makeshift conditions that often ended in contaminations . A few of thousand dollars later, and I've got a sterile, comfortable workplace. The setback of contamination was always a drain on the fun of the hobby for me, especially when it came down to hoping that I didn't stir up too much air in the glovebox, or when attempting to do anything with liquids at all in a small glove box. I want others to be able to enjoy mycology as much as I do, and having a live culture gives much more power when inoculating then trying to start from spores that are hopefully sterile. Kind Regards, David.