Novel MYOC gene mutation, Phe369Leu, in Japanese patients with primary open-angle glaucoma detected by denaturing high-performance liquid chromatography.
OBJECTIVE
To screen for mutations in the MYOC gene in Japanese patients with primary open-angle glaucoma (POAG) using denaturing high-performance liquid chromatography (DHPLC).


METHODS
Blood samples were collected from 171 patients with POAG and 100 controls from seven institutions in Japan. For high-throughput analysis, seven exonic regions were amplified by polymerase chain reaction using DNA pooled from three patients; each DNA pool was then analyzed chromatographically. For analysis of a small number of samples, 7 exonic regions were amplified separately but simultaneously with annealing at 58 degrees C in each patient and then chromatographed, using 7 wells of the same 96-well plate per sample. When chromatographic patterns were abnormal by either method, the PCR products of the individual samples were sequenced.


RESULTS
Four glaucoma-causing mutations were identified in five POAG patients (2.9%). One missense mutation, Phe369Leu, is new; and three others, Ile360Asn, Ala363Thr, and Thr448Pro, have been reported in Japanese patients. Phe369Leu was associated with adult onset POAG.


CONCLUSIONS
Mutations in the MYOC gene were demonstrated chromatographically in 2.9% of our Japanese POAG patients. The use of pooled DNAs with DHPLC analysis is a time- and labor-saving technique. All mutations detected appear to be specific to Japanese patients.
Ishikawa
Karin
K
Department of Ophthalmology, Keio University School of Medicine, Tokyo, Japan.
Funayama
Tomoyo
T
Ohtake
Yuichiro
Y
Tanino
Tomihiko
T
Kurosaka
Daijiro
D
Suzuki
Kotaro
K
Ideta
Hidenao
H
Fujimaki
Takuro
T
Tanihara
Hidenobu
H
Asaoka
Ryo
R
Naoi
Nobuhisa
N
Yasuda
Noriko
N
Iwata
Takeshi
T
Mashima
Yukihiko
Y
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000595:Amino Acid Sequence; D044466:Asian People / Q000235:genetics*; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003598:Cytoskeletal Proteins; D004247:DNA; D005136:Eye Proteins / Q000235:genetics; D005820:Genetic Testing; D005902:Glaucoma, Open-Angle / Q000235:genetics* / Q000473:pathology / Q000503:physiopathology; D006023:Glycoproteins / Q000235:genetics; D006801:Humans; D007930:Leucine; D008875:Middle Aged; D020125:Mutation, Missense; D009898:Optic Disk / Q000473:pathology; D010649:Phenylalanine; D016133:Polymerase Chain Reaction / Q000379:methods; D014794:Visual Fields; D000099179:Myocilin
D003598:Cytoskeletal Proteins; D004247:DNA; D005136:Eye Proteins; D006023:Glycoproteins; D007930:Leucine; D010649:Phenylalanine; D000099179:Myocilin