ABSTRACT
The premise of TA cloning is that certain thermostable polymerases add a single deoxyadenosine to the 3' end of amplified fragments in a template-independent fashion. Ligation of the polymerase chain reaction (PCR) product into a vector is more efficient than blunt-end cloning, but much less efficient than sticky-end. The restriction enzyme sequences should not be contained within the DNA segment to be cloned but must be within the cloning site of the appropriate vector. The chapter discusses a PCR-based method that can be used to obtain new information on the 5' and 3' end of the known cDNA sequence. This method has been termed rapid amplification of cDNA ends and differs from library screening and other methods to obtain full-length sequences in that it may be performed in a relatively short period of time.
