Comparative evaluation of 5'-end-sequence quality of clones in CAP trapper and other full-length-cDNA libraries.
To enhance the usefulness of the laboratory mouse and to facilitate the rapid assay of gene functions we have been collecting the entire set of mouse full-length cDNA by one-pass sequencing. To collect full-length cDNA clones efficiently, it is critical to construct high-quality cDNA libraries. In recent years, we have been developing a way to construct full-length cDNA libraries by using biotinylation of the cap structure (the 'CAP-trapper' method) coupled with treatment to increase reverse transcriptase efficiency at high temperature by the addition of trehalose. In this paper we report our evaluation of the quality of CAP trapper and a number of other full-length cDNA libraries, including the results of 5' end analysis of clones in CAP trapper and the other libraries. We used a procedure that compared the 5'-ends of cDNA clones with those of genes in the public databases. Our analysis showed that 63% of cDNA clones in CAP trapper libraries had sequences that were either the same length as those of equivalent genes in the public database or 5'-extended, and that 90% of these clones maintained their coding sequences. These results indicate that the CAP trapper library is a promising tool for collecting full-length cDNA in large-scale projects. Comparison of the quality of CAP trapper with that of other full-length-cDNA libraries confirmed the value of these libraries.
Laboratory for Genome Exploration Research Project, Genomic Sciences Center and Genome Science Laboratory, RIKEN Tsukuba Institute, 3-1-1 Koyadai, Tsukuba, Ibaraki 305-0074, Japan.
Y
Y
Sugahara
P
P
Carninci
M
M
Itoh
K
K
Shibata
H
H
Konno
T
T
Endo
M
M
Muramatsu
Y
Y
Hayashizaki
D000465:Algorithms; D000818:Animals; D001483:Base Sequence; D018076:DNA, Complementary / Q000235:genetics*; D016208:Databases, Factual; D020224:Expressed Sequence Tags; D015723:Gene Library; D051379:Mice; D008969:Molecular Sequence Data; D012315:RNA Caps / Q000235:genetics*; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid; D012984:Software
D018076:DNA, Complementary; D012315:RNA Caps