The original needs to be well attached to the joists...if it isn't when you have the top layer off, fix that.
The top ply layer should cross the joists, not be parallel with them, for maximum strength. Screw into the subfloor, not the joists - this helps decouple the two layers. There's a good article on this over in the 'Liberry' (sic) at www.johnbridge.com where they specialize in tiling. Even if you aren't going to tile, it will tell you the proper way to minimize deflection and therefore squeaks.