Construction of a normalized directionally cloned cDNA library from adult heart and analysis of 3040 clones by partial sequencing.
Large-scale sequencing of clones from cDNA libraries derived from specific tissues is a rapid and efficient way of discovering novel genes expressed in those tissues. However, because the heart is continually contracting and relaxing, it strongly expresses muscle-contractile genes and/or mitochondrial genes, a bias that reduces the efficiency of this method. To improve the efficiency of identifying novel genes expressed in the heart, we constructed a normalized directionally cloned cDNA library from adult heart and partially sequenced 3040 clones. Comparisons of these sequence data with known DNA sequences in the database revealed that 57.1% of the clones matched human genes already known, 23.4% were identical or almost identical to human expressed sequence tags (ESTs), 14.2% bore no significant homology to any sequences in the database, and 1.2% represented repetitive sequences. The remaining 4.1% showed some homology with known genes, and Northern blot analysis of several clones in this category revealed that most of them were expressed mainly in the heart and skeletal muscle. After redundancy was excluded, the 3040 clones accounted for 1395 distinctive ESTs, 446 of which exhibited no match to any known sequence. Our results suggest that our normalized library is less redundant than standard libraries and is a useful resource for cataloging genes expressed in the heart.
Tanaka
T
T
Laboratory of Molecular Medicine, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo, 108, Japan.
Ogiwara
A
A
Uchiyama
I
I
Takagi
T
T
Yazaki
Y
Y
Nakamura
Y
Y
D000328:Adult; D001483:Base Sequence; D003001:Cloning, Molecular; D015870:Gene Expression; D015723:Gene Library; D005796:Genes; D006801:Humans; D008969:Molecular Sequence Data; D009206:Myocardium / Q000737:chemistry*; D012333:RNA, Messenger / Q000235:genetics* / Q000302:isolation & purification; D012091:Repetitive Sequences, Nucleic Acid; D017422:Sequence Analysis, DNA / Q000379:methods*; D012689:Sequence Homology, Nucleic Acid
D012333:RNA, Messenger