Identification of genes specifically expressed by human Müller cells by use of subtractive hybridization.
OBJECTIVE
Müller cells are the predominant type of glial cells in the retina. They play a critical role in the retina. The purpose of this study was to generate a profile of the genes specifically expressed by human retinal Müller cells and to identify genes that may be responsible for retinal diseases.


METHODS
Subtractive hybridization is a method process by which two populations of mRNA are compared in order to obtain clones of genes expressed in one population but not in the other. A cDNA subtraction library was constructed using RNA isolated from human Müller cells and human astrocytes. PCR-select differential screening was used to further verify the differentially expressed cDNA clones. Positive clones were sequenced and analyzed using the NCBI BLASTN program to identify sequence homologies.


RESULTS
We identified 194 clones specifically expressed in human Müller cells. Among these clones, 102 corresponded to known human genes. Of the remaining 94 clones, 75 corresponded to expressed sequence tags or genomic clones and 19 transcripts did not match with any sequence in databases, and are possibly novel genes.


CONCLUSIONS
The analysis of the subtraction library revealed genes that are specifically expressed by human Müller cells. Some of these genes are unidentified, novel genes that are specific to Müller cells as determined by RT-PCR and Northern blot analyses. These novel genes thus represent candidate genes for retinal diseases.
Lupien
Caroline B
CB
Unité de Recherche en Ophtalmologie, Centre de recherche du CHUQ, Pavillon CHUL, , Ste-Foy, Québec, Canada.
Salesse
Christian
C
D000293:Adolescent; D000328:Adult; D001253:Astrocytes / Q000378:metabolism; D015152:Blotting, Northern; D002478:Cells, Cultured; D002648:Child; D002675:Child, Preschool; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D020224:Expressed Sequence Tags; D015870:Gene Expression; D020869:Gene Expression Profiling; D015723:Gene Library; D015894:Genome, Human; D006801:Humans; D009693:Nucleic Acid Hybridization / Q000379:methods*; D012333:RNA, Messenger / Q000378:metabolism; D012160:Retina / Q000166:cytology* / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D018076:DNA, Complementary; D012333:RNA, Messenger