The phage DNA is isolating by norgen DNA isolation kit and the titre is around 1012.During isolation some host DNA are bound with phage proteins which are not removing by Dnase I digestion.This can not be removed by protenase K or heat treatment as phage coat will be destroyed. Can anyone suggest how this problem can be overcome?
We usually use the sodium chloride at a concentration of 0.5 M. The high salt concentration inhibits the ionic interaction of DNA and proteins. DNase activity while decreasing, but still sufficient for the hydrolysis of bacterial DNA.
You can also use a lower salt concentration of 0.3 M or 0.4 M. Approximately 90% of E.coli proteins usually eluted from the P11-phosphocellulose column at a salt concentration of about 0.4 M. Classic at Arthur Kornberg P11 is an analogue of the DNA.
Protection of bacterial DNA from DNase carry more bacterial proteins whose concentration is high in the lysate than phage proteins.
The classic way to do this would be to purify your phage by ultracentrifugation on a cesium chloride density gradient. But you need a lot of phage - a 500 ml culture is the usual starting point - and you need an ultracentrifuge.
Incidentally, it's worth noting that some phages may pack chromosomal DNA into some of their capsids by mistake. This is the basis of general transduction. However, it will only represent a small percentage of the total phage DNA. However, if this is your problem, there is virtually nothing you can do about it.
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