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Large-scale sequencing of the rabbit corneal endothelial cDNA library
T Fujimaki1, Y Hotta, H Sakuma
1Department of Ophthalmology, Juntendo University School of Medicine, Tokyo, Japan. fujimaki@med.juntendo.ac.jp
Purpose:
This study sought to identify novel or active genes in corneal endothelial cells with description of the gene-expression profile.
Methods:
We performed the single-pass sequencing of 1,000 clones from a rabbit corneal endothelial cDNA library. Inserts of the library were amplified by polymerase chain reaction (PCR), sequenced, and compared with several databases. We used four database similarity search programs: FASTA, BLASTN, TBLASTX, and BLASTX.
Results:
Of the sequences generated, 618 (61.8%) showed sequence homology with known genes, whereas 192 (19.2%) matched previous reported expression sequence tags (ESTs) and 174 (17.4%) showed no sequence similarity. Among the homologous clones to known genes are collagen type VIII, secreted protein acidic and rich in cysteine (SPARC), lysyl oxidase, phosphatidylcholine-2-acylhydrolase, and thrombospondin. Several matched ESTs, and no matched clones that showed high frequency were also detected.
Conclusion:
Large-scale sequencing can be useful in obtaining a profile of the active genes. Several ESTs showed relatively frequent expression, suggesting that these genes may have important functions in the corneal endothelium.