Computational protocol: Genomic and Physiological Analysis of Carbon Storage in the Verrucomicrobial Methanotroph “Ca. Methylacidiphilum Fumariolicum” SolV

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Protocol publication

[…] Genes encoding proteins involved in glycogen metabolism were identified in the available draft genome of strain SolV by Blast searches (Khadem et al., ) which also showed amino acid identities to homologous proteins. Representative reference glgA sequences, encoding the glycogen synthase, were obtained from GenBank and aligned using the MUSCLE aligner in MEGA 5.0 (Tamura et al., ). Phylogenetic trees were calculated using the neighbor-joining method with 1000 bootstraps to infer the evolutionary relationship. Positions containing alignment gaps and missing data were eliminated only in pairwise sequence comparisons (pairwise deletion option). The Dayhoff-matrix based method was used to compute the evolutionary distances. For transcriptome analysis RNA was extracted from exponentially growing cells as described before (Khadem et al., ). After synthesis of cDNA, single-end Illumina sequencing was performed and transcription analysis was performed using the RNA-Seq Analysis tool from the CLC Genomic Workbench software (version 5.0, CLC-Bio, Aarhus, Denmark) and values are expressed as reads per Kilobase of exon model per million mapped reads (RPKM; Mortazavi et al., ). Raw RNA-seq sequence data are available from the GEO depository under accession number GSE40528. […]

Pipeline specifications

Software tools MUSCLE, MEGA
Applications Phylogenetics, RNA-seq analysis, Nucleotide sequence alignment
Organisms Methylacidiphilum infernorum
Chemicals Calcium, Carbon, Carbon Dioxide, Methane, Nitrogen