Computational protocol: Transhydrogenase and Growth Substrate Influence Lipid Hydrogen Isotope Ratios in Desulfovibrio alaskensis G20

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Protocol publication

[…] We constructed a gene tree of nfnAB sequences by retrieving data from two public repositories of annotated genomes: the SEED () and IMG (). For this study, we restricted sequences to those also found in the subsystem “NADH-dependent reduced ferredoxin:NADP+ oxidoreductase” in the SEED (197 total sequences) and manually refined the list to sequences from known sulfate reducers, methanogens, and other anaerobes. Sequences from organisms known to contain nfnAB but not present in the SEED were added manually using a targeted amino acid BLAST search of that organism’s genome in the IMG database. This resulted in a total of 105 nfnAB sequences closely related to that of D. alaskensis G20 using established criteria (; ), with >40% amino acid identity and BLASTP percent identities ranging from 45 to 77%. D. alaskensis G20 itself has two copies of nfnAB that share 85% amino acid identity. A multiple sequence alignment of the 105 putative nfnAB sequences was created using MUSCLE () and checked manually using AliView (). Alignments in FASTA format are available to download in the Supplementary Materials. Pairwise distances for construction of a phylogenetic tree were calculated using the RAxML maximum likelihood algorithm () with the program raxmlGUI (). The tree itself was generated using the Interactive Tree of Life software (). […]

Pipeline specifications

Software tools BLASTP, MUSCLE, AliView, RAxML, raxmlGUI, iTOL
Applications Phylogenetics, Nucleotide sequence alignment
Organisms Desulfovibrio alaskensis G20
Chemicals Fatty Acids, Fatty Acids, Unsaturated, Hydrogen, NADP, Sulfur, Pyruvic Acid