PMID: 11309335Apr 20, 2001Paper

Amplification and expression of splice variants of the gene encoding the P450 cytochrome 25-hydroxyvitamin D(3) 1,alpha-hydroxylase (CYP 27B1) in human malignant glioma

Clinical Cancer Research : an Official Journal of the American Association for Cancer Research
R MaasEckart Meese

Abstract

Recently, we reported the isolation of six novel genes termed glioma-amplified sequences (GASs) from the glioblastoma cell line TX3868 using microdissected mediated cDNA capture (U. Fischer et al., HUM: MOL: GENET:, 5: 595-600, 1996). The aim of this study was to further characterize the gene GAS89. To determine the amplification frequency, we performed comparative PCR studies and Southern blot hybridization experiments. To identify full-length clones of GAS89 we screened a HybriZAP library. Reverse transcription-PCR was performed to isolate splice variants and to determine expression levels. We identified for the gene GAS89 an amplification frequency of 25% in 28 examined glioblastoma multiforme samples. Screening a HybriZAP library, we isolated an incomplete gene sequence showing identity with the gene for 25-hydroxyvitamin D(3) 1,alpha-hydroxylase. Different full-length clones were then isolated using PCR primers chosen from the 3'- and 5'-untranslated regions. As determined by sequencing, the clones represent various splice variants of the 25-hydroxyvitamin D(3) 1,alpha-hydroxylase gene. The clones encode truncated proteins but also one potentially functional enzyme variant. Reverse transcription-PCR studies revealed overex...Continue Reading

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