Balanced-size and long-size cloning of full-length, cap-trapped cDNAs into vectors of the novel λ-FLC family allows enhanced gene discovery rate and functional analysis

We have developed a new class of cloning vectors: lambda -full-length cDNA (lambda -FLC) cloning vectors. These vectors can be bulk-excised for preparing full-length cDNA libraries in which a high proportion of the plasmids carry large inserts that can be transferred into other (for example, functional) vectors. Unlike other cloning vectors, lambda -FLC vectors accommodate a broad range of sizes of eukaryotic cDNA inserts because they contain size balancers. Further, the main protocol we use for direct bulk excision of plasmids is mediated by a Cre-lox system and is apparently free of size bias. The average size of the inserts from excised plasmid cDNA libraries was 2.9 kb for standard and 6.9 kb for size-selected cDNA. The average insert size of the full-length cDNA libraries was correlated to the rate of new gene discovery, suggesting that effectively cloning rarely expressed mRNAs requires vectors that can accommodate large inserts from a variety of sources. Part of the vectors are also suitable for bulk transfer of inserts into various functional vectors.