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Protocol publication

[…] by Ambion. Amounts of RNA were equalised and cDNA was generated using a standard reverse-transcription protocol using random primers (Promega), Superscript II (Invitrogen), First Strand Buffer (Invitrogen) and dNTPs (Promega). PCR using primers designed to NDUFV3 (ENSE00001436763 NDUFV3f1 5′-TGTTTGCTGCGGCAAGGAC-3′ NDUFV3f2 5′- AGCTGCTGTGGCCCTGCTTG-3′) and PCBP3 (ENSE00001682409 PCBP3R1 5′-CTCCCTGATCTCCTTGATCTTG-3′ and ENSE00001303536 PCBP3R1 5′- TCCAGCATGACCACACAGATCTG-3′) were used to check expression of the novel fusion gene ()., Paired-end reads were mapped to a composite reference of the human reference sequence for chromosome 21 (GRCh37) and the mouse genome reference (NCBI m37) using BWA version 0.5.7 (1). Duplicate fragments were marked per library using picard and qualities recalibrated using GATK. SNPs were called using samtoolsSnp. SNP consequences were assigned based on gencode annotation (in Ensembl version 57). SNPs were assigned a dbSNP reference where available. Homozygous SNPs include a splice site mutation in the pseudogene, CR392039 and four non-synonymous Tc1-Hsa21 coding mutations. An amino acid substitution, at a position conserved in mouse and rat, was detected in the steroid co-repressor, NRIP1 that is up-regulated in DS . Non-synonymous changes also occur at conserved sites in the keratin associated protein family genes KRTAP10-10 and KRTAP6-1 that function in hair development , . A mutation in the final coding exon of the testis-expressed gene UMODL1 results in t […]

Pipeline specifications

Software tools BWA, Picard, GATK
Organisms Mus musculus, Homo sapiens