Shearing DNA

[ed]

Ian,


why don't you try nebulizing the DNA ?  someone in this department uses
nebulization very frequently and has his system finely tuned to generate a
median fragment size of about 3 Kb. He uses it for making shotgun
libraries for large scale sequencing. If you are interested, drop me a
line and I can pass this person's e-mail address to you. The procedure is
so foolproof that we used it on an undergraduate molecular biology lab :)


cheers,

Ed

> I've purified some DNA that's larger than 30,000 bp.  I want to shear it
> down to fragments of between 1000-5000 bp.  (For various reasons I want to
> do this mechanically, rather than using restriction ensymes.)  I've been
> trying passing the DNA through a needle, but I seem to be having trouble
> getting the right size out.  For example, passing the DNA through a
> 26-gauge needle 10 times produces DNA that seems smaller than the
> original, but it's still >10 kb.  But passing through a 28-gauge needle 4
> times produces very small fragments, <500 bp.  
> 
> As you can imagine, I don't have an infinite amount of this prep for
> trial-and-error shearing.  Does anyone have a guaranteed, tested and
> consistent procedure?  Or is trial and error the only way to go?