How can I subclone MGC cDNAs from pCMV-SPORT6 into another Gateway vector?
The MGC clones were created by conventional restriction/ligation into the multiple cloning site of the pCMV-SPORT6 vector. The original vector had attB sites and these remain unchanged. The only Gateway reaction available with these clones is BxP; you will need an empty Gateway entry vector with attP sites. This Gateway reaction will yield an entry construct with attL sites (BXP=L). This can then be used to transfer the cDNA to a destination vector with attR (in an LxR=B reaction) to yield the final destination construct with attB sites. So in summary, you will need an empty entry vector and an empty destination vector to subclone into and will have to perform two Gateway reactions. Although this strategy may be cumbersome, PCR errors will be avoided.