These authors contributed equally to this work.
Amplicon-based high-throughput pooled sequencing identifies mutations in CYP7B1 and SPG7 in sporadic spastic paraplegia patients
Article first published online: 13 JUN 2011
© 2011 John Wiley & Sons A/S
Special Issue: Exome Sequencing
Volume 80, Issue 2, pages 148–160, August 2011
How to Cite
Schlipf, N., Schüle, R., Klimpe, S., Karle, K., Synofzik, M., Schicks, J., Riess, O., Schöls, L. and Bauer, P. (2011), Amplicon-based high-throughput pooled sequencing identifies mutations in CYP7B1 and SPG7 in sporadic spastic paraplegia patients. Clinical Genetics, 80: 148–160. doi: 10.1111/j.1399-0004.2011.01715.x
- Issue published online: 12 JUL 2011
- Article first published online: 13 JUN 2011
- Accepted manuscript online: 27 MAY 2011 09:49AM EST
- Received 10 May 2011, revised and accepted for publication 24 May 2011
- amplicon-based next-generation sequencing;
- hereditary spastic paraplegia;
Schlipf NA, Schüle R, Klimpe S, Karle KN, Synofzik M, Schicks J, Riess O, Schöls L, Bauer P. Amplicon-based high-throughput pooled sequencing identifies mutations in CYP7B1 and SPG7 in sporadic spastic paraplegia patients.
Hereditary spastic paraplegia (HSP) is a neurodegenerative disorder defined clinically by progressive lower limb spasticity and weakness. HSP is a genetically highly heterogeneous condition with at least 46 gene loci identified so far, involving X-linked, autosomal recessive (AR) and autosomal dominant inheritance. For correct diagnosis, molecular testing is essential because clinical parameters by themselves are not reliable to differentiate HSP forms. The purpose of this study was to establish amplicon-based high-throughput genotyping for AR-HSP. A sample of 187 index cases with apparently sporadic or recessive spastic paraplegia were analyzed by applying an array-based amplification strategy. Amplicon libraries of the CYP7B1–(SPG5) and SPG7–gene were generated followed by a pooled next-generation sequencing (NGS) approach. We identified three SPG5 and seven SPG7 patients. All had one homozygous or two heterozygous mutations. In total, 20 distinct mutations (CYP7B1,n = 4and SPG7,n = 16) including two novel CYP7B1 mutations (p.G51R and p.E211KfsX3) and eight novel SPG7 mutations (p.Leu8delinsLeuLeu, p.W29X, p.R139X, p.R247X, p.G344D, p.Leu346_Leu347ins11, p.R398X and p.R398Q) were detected by this comprehensive genetic testing. Our study illustrates how amplicon-based NGS can be used as an efficient tool to study genotypes and mutations in large patient cohorts and complex phenotypes.