Link to this post | posted 05 Jul, 2017 21:47

scaruso

Joyce, I use a slightly different protocol that I found online and liked, but it's only different in the details, the idea is the same. I would use whichever works best, or easiest, for you. For mine, you will want some cotton/glass/fiberglass wool, as above, which you can put in a funnel and cover in foil for autoclaving. It makes a nice long term dispenser that you can re-cover with the foil. 1 - Spread 100 uL of a growing culture on two plates and incubate ~4-5 days at 30ºC. 2 - Assemble the filter - place a sterile 10 mL syringe in a sterile 50 mL conical tube and remove plunger, use sterile forceps to push wool into bottom of syringe. 3 - Add ~5 mL sterile ddH2O to the each confluent plate. 4 - Using sterile cotton applicators (or spreaders as Lee uses), gently rub spores off surface of plates 5 - Transfer the liquid using a filtered pipette tip to the 10 mL syringe, and push through the wool using the plunger 6 - Remove the syringe, replace cap, and spin the conical tube 5' @ ~5kg 7 - Carefully remove supernatant, then resuspend the pellet in the minimum volume of sterile 20% glycerol (~500 uL) 8 - Transfer to a 2 mL screw cap tube and store at -20ºC. I'm not sure what lab the original came from, but a longer version of the protocol can be found here: Protocols - Spore Prep Edited 05 Jul, 2017 21:49

Link to this post | posted 29 Dec, 2016 22:43
scaruso	I see that the web phamerator can make maps, and understand that other features are coming. Is there a way to have it access alternate databases that I don't see (like Bacillus phages), of is that one of the not yet ready for prime time features? Thanks! steve

Link to this post | posted 24 May, 2016 16:02
scaruso	What a tragic accident. David will be missed, and I will miss the excitement and energy he brought with him to each new challenge. Edited 24 May, 2016 16:02

Link to this post | posted 16 Nov, 2015 20:35
scaruso	Hello, Is there a link we can provide to our OIT depts or do I need to give them my log-in information? Steve

Link to this post | posted 16 Sep, 2015 14:05

scaruso

Hello all, I have started working with Strep. griseus and would like to solicit some advice. Lee generously provided me with a lot of information, but, honestly, these are critters that I have seen for the first time. I am growing them in nutrient broth (NB) + PEG to prevent clumping then using NB + supplements described by Lee for enrichments, and nutrient agar with the same supplements for plating. I have a culture being maintained at 30-deg w/ shaking in NB+PEG which I am sub-culturing for students, and I am starting it fresh on Fridays from a spore stock I created. So, my issues. Lawns with Strep. are not dense since TA is not used, so plaques are really tough to see. So far, those that the students have tested have proven not to be such. So, no phage so far, which is discouraging, of course. The cultures are pretty dense, but I am wondering how/if I can make thicker lawns or increase the likelihood of picking up a phage. Any thoughts are appreciated. Steve