Henry Swanson said:

This is an exerpt from the post I linked. It's worked well for me, which is why I was sharing. Different strokes (no pun intended) for different folks

c10h12n2o said:

Working with Spores:
since we fruit in open air, we can assume that spores are dirty. if someone sends you a spore print, you can assume it is going to have contams on it in addition to the spores you want to cultivate. this does not mean you got a bad print, it means you got a NORMAL print. the same thing applies to syringes. for this reason, when inoculating something with spores, it is a good idea to use as little as possible, since we only need a few spores and the more solution/spores we use, the more contams will be present as well. i recommend people never putting spores directly to grain, since it is so easy to culture the contams

a sterile jar of grain is basically the microbiological jackpot for anything that can digest it (fungi, bacteria, etc), so we need to take care to reduce the chances of culturing the wrong microorganisms. basically, ALWAYS germinate your spores first on agar before applying them to grain/lc/etc.

by using streaking techniques, you can easily clean up even the dirtiest of prints or spore syringes. you basically are using a loop to spread spores VERY thin across the surface of agar, creating various zones with different concentrations of spores. this is extremely useful for cleaning up a print, since you spread the contams thin while you spread the spores. this is also a great way to obtain isolates about 100 transfers sooner than you would get from a blob of spores, since the later zones have very few strains present in each colony. by doing one of these serial dilution style streaks, you end up with hundreds of colonies to choose from, of various genetic diversity, which gives you lots of chances to select a vigorous, organized culture

Quote:

In microbiology, streaking is a technique used to isolate a pure strain from a single species of microorganism, often bacteria(fungi in our case). Samples can then be taken from the resulting colonies and a microbiological culture can be grown on a new plate so that the organism can be identified, studied, or tested.

Streaking is rapid and ideally a simple process of isolation dilution. The technique is done by diluting a comparatively large concentration of bacteria to a smaller concentration. The decrease of bacteria should show that colonies are sufficiently spread apart to effect the separation of the different types of microbes. Streaking is done using a sterile tool, such as a cotton swab or commonly an inoculation loop. Aseptic techniques are used to maintain microbiological cultures and to prevent contamination of the growth medium.There are many different types of methods used to streak a plate. Picking a technique is a matter of individual preference and can also depend on how large the number of microbes the sample contains.

The three-phase streaking pattern, known as the T-Streak, is recommended for beginners.The streaking is done using a sterile tool, such as a cotton swab or commonly an inoculation loop. The inoculation loop is first sterilized by passing it through a flame. When the loop is cool, it is dipped into an inoculum such as a broth or patient specimen containing many species of bacteria. The inoculation loop is then dragged across the surface of the agar back and forth in a zigzag motion until approximately 30% of the plate has been covered. The loop then is re-sterilized and the plate is turned 90 degrees. Starting in the previously streaked section, the loop is dragged through it two to three times continuing the zigzag pattern. The procedure is then repeated once more being cautious to not touch the previously streaked sectors. Each time the loop gathers fewer and fewer bacteria until it gathers just single bacterial cells that can grow into a colony.The plate should show the heaviest growth in the first section. The second section will have less growth and a few isolated colonies, while the final section will have the least amount of growth and many isolated colonies.

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examples of streaked plates. different labs use different standards for streaking. colonies thin as the dish rotates counter clockwise

the way i like to do it is very similar to what is described above. basically, take your loop (bod's DIY loop tek) and scrape off a small amount of spores from a print (or drip a drop from a syringe directly onto the loop) and streak a side of the plate like below. then flame the loop again, DONT SCRAPE MORE SPORES, rotate the dish, and drag the loop from the area you streaked previously to streak zone 2, then flame and repeat

this is the streaking technique i like to use. it is tremendously handy for cleaning up spore prints, isolation projects, and even cleaning up old LCs

if you are using a spore syringe as your starting point (or a LC you are trying to refresh), you want to be super careful not to add any additional liquid to the plate, otherwise it will roll around and distort the streaking pattern. so make sure you dont drip over the plate, you shouldnt be putting any more liquid into the plate than what sticks to the loop

Conclusion:

so that is pretty much it!! a bit lengthy, but i think i covered most of what i wanted to cover for my post #1000. please let me know if yall have any questions or need clarification about anything

i also welcome any additional tips & tricks, feedback, criticism, discussion, debate, haters (), or critical thinking

thanks everyone for everything yall have done to make this place such an incredible resource. im so grateful to live in an age where there is an online community to collaborate with on our hobbies

warm regards,
c10

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leproos said:

The fluffy white at the bottom of the stems are pretty big is that normal or what does it say?

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BeefSupremeJr said:

if thats the first to pin you might stil get the expression you want later

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BeefSupremeJr said:

also im not sure why that pyrex should have broken but im not sure about going from the microwave into a bath of relatively cold water or from the fridge straight to high setting on a microwave.  seems like a triathalon for the poor bottle.  its been years since i used a microwave to heat up agar.  my microwave is a pos nobrand i bought at a yard sale for 10$ years ago and i keep around mostly as a joke.  it takes two full minutes to heat up a cup of coffee .

anyway glad youre alright bud you could have been burnt pretty good.  at the end of the day i guess i think its still best to not microwave agar if you can avoid it.  bod always did it and i remember him saying to do it slowly so i have to wonder if he popped a couple media bottles himself..

ive also exploded some casarole dishes coming home drunk trying to get a midnight snack impatiently.

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Quote:

callmehippie said:
For sure. I use this technique as well and also link it to lots of people.

I actually was just telling Gilmour Hendrix about it the other day. Lol

Somtimes i just  like simplify it and just do the zigzag down the middle just cuz that the other way  you show is a tiny bit more complicated.

You've got to torch the Loop multiple times and remove the dish lid multiple times if you're not working in flow. Not so much noob friendly I guess.

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BeefSupremeJr said:

qq I ever tell you guys I dont believe in streaking?

yeah i think its not actually helpful in this hobby.  i mean why reduce the genetic material to work with? i think its pointless in our circumstsnce and doesnt accomplish anything a few transfers couldnt anyway.  its just fancy. 

i mean read anyones process and theyll explain how its done and at the most theyll explain how it helps dilute and isolate cultures.  but why?  WHY do we want that?

i want to go to fruit with as much genetic material as possible.  it youre playing the lottery, does it not make sense to have as many tickets as possible? i mean i also think playing lottery is kindof stupid but mushroom pheno hunting is a litlte more like playng slots.  okay, does it not make sense to play as many machines as possible?

well? does it?

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RoscoeReturns said:

I don’t give a shit about genetic diversity and pheno hunting. I want to spread out my spores so if there is contamination I have a better chance of finding a clean spot to transfer away from/less chance of having multiple organisms growing on top of one another. I think for the majority of people on here asking questions that is probably a more important goal than taking as much genetic diversity to grain.

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