Hello,
I am looking for alternatives or cheaper materials to isolate out MS2 from E. coli following lysis via chloroform. Some literature I have reviewed uses a .45 um nanopore filter following lysis. From what I have seen, these tend to be fairly expensive for our budget. Does anyone have any other suggestions?
One possible approach I have seen is to use a .45um syringe (cheaper) filter. Would this be appropriate? And would this option be as simple as I think: use the plunger to pressure the lysis solution through the filter with a collection container on the other end?
Yes in principle this could work but blockages could be a problem
As Stuart has mentioned, syringe filtration is good for small volumes, as the membrane gets clogged. For larger volumes you can try tangential flow filtration system (TFF). They are available from lab scale to large scale.
Any sterile 0.45 or 0.22 micron filter should be fine to sterilize your lysate and remove the viable E. coli cells. However if you have removed most of the cells and cell debris by centrifugation then just adding a few drops of chloroform to the tube and incubating for a few minutes at 37-43C will also kill any remaining viable cells. You can always test for sterility by spotting 10ul of your lysate on an LB plate and see if anything grows.
I don't remember how heat stable MS2 is, but when I worked with M13 I would often sterilize by incubating at 55-65C for 10 min (M13 is very heat stable).
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