I think it really depends on your mobo. I've used the bottome PCI-e slot before as they cooler air is towards the bottom of your case, so I figured putting my video card as close to that as possible would help it run cooler. Worked fine.
But again, think it depends on your mobo. Give it a go. It'll either work or it wont. One thing to check is that most mobo's give you the option to choose which video to boot from; i.e PCI-e 1 --> PCI-e 2 --->PCI-e 3, etc. If you're using PCI-e 2 make sure that's where the system looks first for a video output.