Expressed sequence tag analysis of human retina for the NEIBank Project: retbindin, an abundant, novel retinal cDNA and alternative splicing of other retina-preferred gene transcripts.
OBJECTIVE
Expressed sequence tag (EST) analysis was performed on un-normalized, unamplified cDNA libraries constructed from adult human retina to examine the expression profile of the tissue and to contribute resources for functional genomics studies.


METHODS
Two size fractionated cDNA libraries (designated hd and he) were constructed from human retina RNA. Clones were randomly selected for sequencing and analyzed using the bioinformatics program GRIST (GRouping and Identification of Sequence Tags). PCR, Northern blotting and other techniques have been used to examine selected novel transcripts.


RESULTS
After informatics analysis, 2200 retina cDNAs yield 1254 unique clusters, potentially representing individual genes. Opsin is the most abundant transcript and other retina transcripts are prominently represented. One abundant cluster of cDNAs encodes retbindin, a novel, retina preferred transcript which has sequence similarity to riboflavin binding proteins and whose gene is on chromosome 19. Variant transcripts of known retina genes are also observed, including an alternative exon in the coding sequence of the transcription factor NRL and a skipped coding sequence exon in the phosphodiesterase gammasubunit (PDE6G).


CONCLUSIONS
The new retina cDNA libraries compare favorably in quality with those already represented in public databases. They are rich in retina specific sequences and include abundant cDNAs for a novel protein, retbindin. The function of retbindin remains to be determined, but it is a candidate for flavinoid or carotenoid binding. Analysis of multiple clones for highly expressed retina genes reveals several alternative splice variants in both coding and noncoding sequences which may have functional significance. The validated set of retina cDNAs will contribute to a nonredundant set for microarray construction.
Wistow
Graeme
G
Section on Molecular Structure and Function, National Eye Institute, National Institutes of Health, Bethesda, MD 20892-2740, USA. graeme@helix.nih.gov
Bernstein
Steven L
SL
Wyatt
M Keith
MK
Ray
Sugata
S
Behal
Amita
A
Touchman
Jeffrey W
JW
Bouffard
Gerald
G
Smith
Don
D
Peterson
Katherine
K
D000368:Aged; D000369:Aged, 80 and over; D017398:Alternative Splicing / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D018076:DNA, Complementary / Q000032:analysis*; D020224:Expressed Sequence Tags; D005136:Eye Proteins / Q000235:genetics* / Q000378:metabolism; D020869:Gene Expression Profiling; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D009316:National Institutes of Health (U.S.); D020411:Oligonucleotide Array Sequence Analysis; D009885:Ophthalmology; D016133:Polymerase Chain Reaction; D012160:Retina / Q000378:metabolism*; D014481:United States
D018076:DNA, Complementary; D005136:Eye Proteins; C459679:RTBDN protein, human