Fine expression profiling of full-length transcripts using a size-unbiased cDNA library prepared with the vector-capping method.
Recently, we have developed a vector-capping method for constructing a full-length cDNA library. In the present study, we performed in-depth analysis of the vector-capped cDNA library prepared from a single type of cell. As a result of single-pass sequencing analysis of 24,000 clones randomly isolated from the unamplified library, we identified 19,951 full-length cDNA clones whose intactness was confirmed by the presence of an additional G at their 5' end. The full-length cDNA content was >95%. Mapping these sequences to the human genome, we identified 4,513 transcriptional units that include 36 antisense transcripts against known genes. Comparison of the frequencies of abundant clones showed that the expression profiles of different libraries, including the distribution of transcriptional start sites (TSSs), were reproducible. The analysis of long-sized cDNAs showed that this library contained many cDNAs with a long-sized insert up to 11,199 bp of golgin B, including multiple slicing variants for filamin A and filamin B. These results suggest that the size-unbiased full-length cDNA library constructed using the vector-capping method will be an ideal resource for fine expression profiling of transcriptional variants with alternative TSSs and alternative splicing.
Oshikawa
Mio
M
Department of Rehabilitation Engineering, Research Institute, National Rehabilitation Center for Persons with Disabilities, 4-1 Namiki, Tokorozawa, Saitama 359-8555, Japan.
Sugai
Yoshiko
Y
Usami
Ron
R
Ohtoko
Kuniyo
K
Toyama
Shigeru
S
Kato
Seishi
S
D002478:Cells, Cultured; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D030561:Databases, Nucleic Acid; D018628:Gene Dosage; D020869:Gene Expression Profiling / Q000379:methods*; D015723:Gene Library; D005822:Genetic Vectors / Q000502:physiology; D006801:Humans; D012315:RNA Caps / Q000737:chemistry / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification; D015203:Reproducibility of Results; D024363:Transcription Initiation Site / Q000502:physiology
D012315:RNA Caps; D012333:RNA, Messenger