Dear Anil,

Try 5 mv constant current in the submerged transfer apparatus, Overnight transfer on PVDF membrane. at 4 degrees. I used to put the apparatus inside the fridge (bottom). It works. 

before transfer equilibrate your membrane in 100% methanol for 5 min, then the transfer buffer.

I hope this should work.

Some of my friends also use biorad's semi dry apparatus and say that it is fast and efficient but I never use that, so If you have one you can try that also, with the manufacturer's protocol, but I think the buffer composition is different for that one.

Good luck

Do you get the same result if you put the gel upside down in the blotting cassette?

Thanks Roopesh and Alejandro. We figured out the problem. It seems someone may have forgotten to add SDS while making 10X running buffer.