Large-scale sequencing of the rabbit corneal endothelial cDNA library.
OBJECTIVE
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.


CONCLUSIONS
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.
Fujimaki
T
T
Department of Ophthalmology, Juntendo University School of Medicine, Tokyo, Japan. fujimaki@med.juntendo.ac.jp
Hotta
Y
Y
Sakuma
H
H
Fujiki
K
K
Kanai
A
A
D000818:Animals; D003094:Collagen / Q000235:genetics; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000032:analysis*; D004728:Endothelium, Corneal / Q000737:chemistry*; D020224:Expressed Sequence Tags; D005136:Eye Proteins / Q000235:genetics*; D005819:Genetic Markers; D015676:Osteonectin / Q000235:genetics; D010713:Phosphatidylcholines / Q000235:genetics; D016133:Polymerase Chain Reaction; D008249:Protein-Lysine 6-Oxidase / Q000235:genetics; D011817:Rabbits; D019699:Thrombospondins / Q000235:genetics
D017931:DNA Primers; D018076:DNA, Complementary; D005136:Eye Proteins; D005819:Genetic Markers; D015676:Osteonectin; D010713:Phosphatidylcholines; D019699:Thrombospondins; D003094:Collagen; D008249:Protein-Lysine 6-Oxidase