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, Classification of Tumours of the Central Nervous System
, All cases were confirmed by STAT6 nuclear staining of tumor cells using an already published protocol, vol.1
, NAB2-STAT6 gene fusion analysis and TERT promoter mutation analysis NAB2-STAT6 gene fusion analysis and TERT promoter mutation analysis was performed on all cases
RNA was extracted from formalin-fixed, paraffin-embedded tissue using RNA-Bee-RNA isolation reagent (Bio-Connect BV, Huissen, the Netherlands). RNA quantity and quality were determined by a NanoDrop measurement ,
The cDNA was tested by the reverse transcription-polymerase chain reaction (RT-PCR) for the HMBS (hydroxymethylbilase synthase) housekeeping gene using the primers forw150, vol.5 ,
, For the detection of the most common NAB2-STAT6 gene fusions, three primers in NAB2 (NM_005967.3): forw 5'-CAAGTAGCCCGAGAGAGCAC-3' (exon 3), forw 5'-CTCCACTGAAGAAGCTGAAAC-3' (exon 4) and forw 5'-CTGTGTGCCTGCGAAGCC-3' (exon 6) were used in combination with three primers in STAT6 (NM_001178078.1): rev 5'-GGGAAAGTCGACATAGAGCC-3' (exon 2), rev 5'-GAGCTGAGCAAGATCCCGG-3' (exon 16) and rev 5'-TTCCACGGTCATCTTGATGG-3
Adapter ligation and barcoding was done using the NEBNext Ultra DNA Library Prep Mastermix Set and NEBNext Multiplex Oligos from New England Biolabs. CLC Cancer Research Workbench software from QIAGEN ® was performed for sequence analysis. Described briefly, the analysis workflow included adapter trimming and read pair merging before mapping to the human reference genome (hg19). Insertions and deletions as well as single nucleotide variants were detected ,
Solitary fibrous tumor -clinicopathologic, immunohistochemical and molecular analysis of 28 cases, Diagnostic pathology, vol.9, p.224, 2014. ,