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BB does silicon lids

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#1 Hippie3



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Posted 20 August 2006 - 09:16 AM

Jar lids are important. The classic PF Tek jar lid uses four nail holes for inoculation and a layer of dry vermiculite to prevent contamination of the substrate. Airborne contaminants are filtered out by the vermiculite and if you inject carefully, the verm slides back to “heal” the injection site when you remove the needle. Dry verm makes an adequate contaminant barrier. We will use the dry verm layer on top of our jars, but we will also use tyvek for filtered air exchange and silicone for self healing injection ports. Ports and Tyvek aren't required, but they certainly don't hurt.
Parts List:
1. Jar lids (metal or plastic)
2. Tyvek (envelopes from the post office)
3. Smallish metal washers (2 for each lid)
4. GE Silicone II caulk (clear)
5. Caulking gun
6. Electric drill
7. 3/16 drill bit
Begin by using the washers to trace out a bunch of circles on the tyvek envelope. Using sharp scissors cut out these circles. You will need two circles for each lid. This is annoyingly time consuming. The process can be sped up by stapling four layers of Tyvek together and cutting all four layers at once.
Now drill five 3/16 inch holes in each lid. Make one hole in the center and four around the outer edges. Don’t go too close to the edges. It is easy to do a large number of lids quickly by stacking them up and drilling them all at once. After drilling, *carefully* remove the sharp metal shavings from the holes. If you aren’t careful, you will cut your fingertips quite badly on the shavings. Use leather gloves and a rat tail file, if you have them. Using plastic lids avoids these sharp metal fragments.
If you do not have access to a drill, you can use a nail to punch the holes..
Lid showing hole positions (top washer already attached).
Cut open the tube of caulk so it will lay a reasonably large bead. Put a large blob of caulk on the top of each of the four outer edge holes. Work the caulk with the gun tip to get it smeared a little around the edges. Don’t go all the way to the lid’s edge with the caulk or the bands won’t sit flat. Once the four holes are “blobbed” on top, flip the lid over and put a smaller blob on the underside. Let the lid dry sitting flat for 24 hours. An empty jar makes a nice support for a lid while it is drying.
Inoculation hole with silicone blob. Note clean edge so band will sit flat.
Inoculation hole underside.
Glue two washers over the center hole (one on top, one on bottom) with silicone. Use the silicone to glue a tyvek circle over each washer (one on top, one on bottom). The point of the washers is to provide a dead air space between the tyvek layers. The dead air space is a physical barrier against any contaminant that might wick through the tyvek by capillary action if it gets wet.
Washer on jar top without tyvek.
Washer and tyvek on jar bottom.
Two lids, one with tyvek and one without.
Let everything dry for another 24 hours before using.
You can inject spores or LC through the 4 silicone blobs and they will close up (heal) when you remove the needle. Do not inject through the tyvek. The double tyvek filter is for air exchange only. If you inject into the silicone blobs in a different spot each time, the ports will last longer. If you inject through the same spot every time, the silicone blob will deteriorate (and subsequently leak) much quicker.
These silicone blobs are now referred to as self healing injection ports.
As soon as a port becomes noticeably deteriorated, scrape it off with a razor blade and apply a fresh one. If you change injection angles, a thick port should be good for hundreds of needle sticks before needing replacement. When using injection ports, sharp (not blunted) needles are preferred.
Using this lid design, you could omit the dry vermiculite layer called for in the classic PF Tek, but you shouldn’t. Tyvek can breathe so well (even through two layers) that a dry spot forms right below the filter. A dry spot on a cake won’t colonize and is a contamination vector after birth. The verm layer will prevent dry spots.
Make at least two lids for liquid culture jars. Liquid culture jars don’t need tyvek vents, just two silicone injection ports.<O:p

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