SMART cDNA synthesis method utilizes specific feature of Moloney murine leukemia virus reverse transcriptase (RT) to add a few non-template deoxynucleotides (mostly C) to the 3' end of a newly synthesized cDNA strand upon reaching the 5' end of the RNA template (Schmidt and Mueller, 1999). When added to the RT reaction, oligonucleotides containing an oligo(riboG) sequence on the 3' end (TS-oligo) form pairs with the deoxycytidine stretch produced by RT. The reverse transcriptase continues replication using the TS-oligo as a template, leading to 3' addition of the complimentary TS-sequence to serve as a universal 5' terminal site for primer annealing during following cDNA amplification (Chenchik et al., 1998; Matz et al., 1999).
SMART-prepared cDNA is full-length-enriched and well suitable for generation of cDNA libraries for various applications (Zhu et al., 2001).
Schematic outline of SMART cDNA synthesis.