Hi Abdullateef,

I suggest:

• If the positive control is also not successful, check competent cells with pUC19 or try to increase the amount of cells that you plated.
• If the positive control works fine, try increasing the incubation time to 18h. Maybe you forgot to treat reactions with proteinase K before attempting transformation or you are not using the correct antibiotic for selection. Are you using the recommended amount of DNA? Maybe your attB-PCR or the linear attB is to long, if it is longer than 5kb try incubating the BP reaction overnight. Or maybe you just made some little mistake designing your att sequences.

Hope it helps. Good luck!!!

Thanks for the useful Tips Brian and Mark

i will try to check the insert/donor amount and update the results 

i still wonder weather it is critical to use the PE buffer or not. i tried to follow the protocol i.e elution of PCR product with PE after size purification with 10% PEG 8000 and alternatively i used the PCR product (in PCR buffer ) and plasmid ( mini prep kit ellution buffer) only 

Here is my update and sorry for lateness. i did size purification using PEG following the protocol and used TOP10 CC and i got tens of colonies. i progressed with my work but still not that convincing efficiency probably due to the PB enzyme i'm using it was sitting in the freezer for quite long time and it is shared by all lab members.

thank you for the helpful tips