There's no performance difference, as they compile to the same bytecode:
Python 2.6.2 (r262:71600, Apr 15 2009, 07:20:39)
>>> import dis
>>> def f(x):
... return x is not None
...
>>> dis.dis(f)
2 0 LOAD_FAST 0 (x)
3 LOAD_CONST 0 (None)
6 COMPARE_OP 9 (is not)
9 RETURN_VALUE
>>> def g(x):
... return not x is None
...
>>> dis.dis(g)
2 0 LOAD_FAST 0 (x)
3 LOAD_CONST 0 (None)
6 COMPARE_OP 9 (is not)
9 RETURN_VALUE
Stylistically, I try to avoid not x is y. Although the compiler will always treat it as not (x is y), a human reader might misunderstand the construct as (not x) is y. If I write x is not y then there is no ambiguity.