Computational protocol: A mutation of EPT1 (SELENOI) underlies a new disorder of Kennedy pathway phospholipid biosynthesis

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Protocol publication

[…] Genomic DNA samples were extracted from peripheral blood following standard protocols. Genome-wide genotyping was carried out using Illumina HumanCytoSNP-12 v2.1SNP arrays. Data output was visualized in Illumina’s GenomeStudio software. Multipoint linkage analysis was performed assuming a recessive mode of inheritance, full penetrance, and a disease allele frequency of 0.0001 using SimWalk2 ().Whole exome sequencing was performed by Otogenetics Corp. using the SureSelect Human All Exon V4 (Agilent Technologies) exome enrichment kit on an Illumina HiSeq2000. Reads were analysed on the DNAnexus platform for exome coverage, SNP/InDel variant calling and quality filtering. The exome sequencing produced 21 907 356 mapped 100 bp paired-end reads, matching 95.92% of targeted sequences adequately covered for variant calling (>10× coverage; mean depth, 38.4×).The identified putative mutations were validated by PCR amplification followed by di-deoxy sequence analysis (Applied Biosystems 3130 DNA Sequencer, Life Technologies). Primer design to amplify the coding exon 5 of the SELENOI/EPT1 gene was done using Primer3 online tool (). Primer sequence specificity was verified by using the UCSC In-Silico PCR tool and their sequences were screened to exclude common single nucleotide polymorphisms. […]

Pipeline specifications

Software tools GenomeStudio, SimWalk2, DNAnexus, Primer3, In-Silico PCR
Applications Miscellaneous, WES analysis, qPCR
Organisms Homo sapiens
Diseases Spastic Paraplegia, Hereditary, Heredodegenerative Disorders, Nervous System
Chemicals Cytidine Diphosphate, Phosphatidylcholines, Ethanolamine