Comparative molecular approaches in Prader-Willi syndrome diagnosis.
Prader-Willi and Angelman syndromes are two distinct neurogenetic disorders caused by chromosomal deletions, uniparental disomy or loss of the imprinted gene expression in the 15q11-q13 region. PWS results from the lack of the paternally expressed gene contribution in the region. The aim of our study was to compare a new molecular approach based on the quantification of the expression of non-imprinted bi-allelic gene (NIPA1 and OCA2) with in house MS-PCR and the MS-MLPA test. Blood samples were collected from 12 patients, clinical criteria positives for Prader-Willi syndrome. DNA and RNA samples were isolated from white blood cells. Epigenetic changes at SNRPN gene locus were evaluated by MS-PCR technique. The expression levels of two non-imprinted genes (NIPA1 and OCA2) were evaluated in qReal-Time PCR, in order to identify type 1 and type 2 deletions. SALSA MS-MLPA kit ME028 was used to detect copy number changes and to analyze CpG islands methylation of the 15q11 region. MS-MLPA test confirmed that 8/12 patients presented different types of deletion at the SNRPN gene level (promoter, introns, and exons) and 4/8 displayed type 1 or type 2 deletion. In children with 15q11-13 deletions, the decreased level of NIPA1and OCA2 gene expression is related to chromosomal abnormality in the investigated area. The deletions were confirmed by MS-MLPA analysis, thus recommending NIPA1 and OCA2 gene expression as an alternate method to investigate deletions.
"Stefan S. Nicolau" Institute of Virology Bucharest, Romania. Electronic address: gnanka30@yahoo.com.
Anca
A
Botezatu
"V.Babes" University of Medicine and Pharmacy Timisoara, Romania.
Maria
M
Puiu
University of Bucharest, Faculty of Biology, Bucharest, Romania.
Natalia
N
Cucu
"Stefan S. Nicolau" Institute of Virology Bucharest, Romania.
Carmen C
CC
Diaconu
"C.I. Parhon" Institute of Endocrinology Bucharest, Romania.
C
C
Badiu
University of Bucharest, Faculty of Biology, Bucharest, Romania.
C
C
Arsene
"Stefan S. Nicolau" Institute of Virology Bucharest, Romania.
Iulia V
IV
Iancu
"Stefan S. Nicolau" Institute of Virology Bucharest, Romania.
Adriana
A
Plesa
"Stefan S. Nicolau" Institute of Virology Bucharest, Romania.
Gabriela
G
Anton
D002648:Child; D002675:Child, Preschool; D002869:Chromosome Aberrations; D002884:Chromosomes, Human, Pair 15 / Q000235:genetics / Q000378:metabolism; D018899:CpG Islands; D019175:DNA Methylation; D005260:Female; D005786:Gene Expression Regulation; D056426:Genetic Loci; D006801:Humans; D007223:Infant; D008297:Male; D008565:Membrane Proteins / Q000096:biosynthesis / Q000235:genetics; D026901:Membrane Transport Proteins / Q000096:biosynthesis / Q000235:genetics; D011218:Prader-Willi Syndrome / Q000175:diagnosis / Q000235:genetics / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D017384:Sequence Deletion
D008565:Membrane Proteins; D026901:Membrane Transport Proteins; C478824:NIPA1 protein, human; C489600:OCA2 protein, human
Genomic imprinting; MLPA analysis; Prader–Willi Syndrome