Identification and characterization of NF1 mutations using single strand conformational polymorphism (SSCP) analysis

Journal Article · Thu Sep 01 04:00:00 EDT 1994 · American Journal of Human Genetics

OSTI ID:134769

Rodenhiser, D I; Hovland, K; Singh, S M ^[1]

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1. Children`s Hospital of Western Ontario (Canada); and others

Neurofibromatosis type 1 (NF1) is one of the most common human genetic disorders with a constellation of cutaneous and skeletal manifestations, intellectual impairment, and an increased risk for a variety of malignancies. The neurofibromin gene is also considered a tumor-suppressor gene since its loss of function is associated with a variety of sporadic cancers in the general population. The NF1 gene has a high spontaneous mutation rate, and while a number of laboratories are involved in a coordinated effort to identify NF1 mutations, only a small number of mutations have been characterized. Despite considerable efforts no high frequency or recurrent mutation has been found. We report the application of single strand conformational polymorphism (SSCP) and heteroduplex analysis on the Phastgel system to identify mutations in the neurofibromin gene. A DNA panel of patients representing 100 families from Ontario, Canada was used to screen fourteen NF1 exons encompassing 30% of the NF1 gene: the 5{prime} exons 1, 17, 24 and the 3{prime} exons 28-33, 39-42 and 49. SSCP and heteroduplex variants were identified in PCR products amplified from 8 exons and mutations were identified in 10% of patients. Three RFLPs also have been identified and three other SSCP variants are being characterized. While most small deletions and insertions form heteroduplexes readily detectable on native gels, our results suggest that the detection of heteroduplexes resulting from point mutations is best facilitated on native Phastgels at low temperature. Our results suggest that as point mutations comprise a significant proportion of NF1 mutations, optimization of the SSCP protocol is critical to ensure the detection of all sequence variants.

• OSTI ID: 134769
• Report Number(s): CONF-941009--
• Journal Information: American Journal of Human Genetics, Journal Name: American Journal of Human Genetics Journal Issue: Suppl.3 Vol. 55; ISSN AJHGAG; ISSN 0002-9297
• Country of Publication: United States
• Language: English

Related Subjects

55 BIOLOGY AND MEDICINE
BASIC STUDIES
DETECTION
EXONS
GENE MUTATIONS
GENES
MENTAL DISORDERS
MUTATION FREQUENCY
NEOPLASMS
ONTARIO
PATIENTS
POLYMERASE CHAIN REACTION
RFLPS
SIZE
SKELETAL DISEASES
SKIN DISEASES
SPONTANEOUS MUTATIONS