Solani PEL-D [AAC49420]87 53Fusarium solani PEL-B [AAA87383] Fusarium …

Solani PEL-D [AAC49420]87 53Fusarium solani PEL-B [AAA87383] Fusarium oxysporum [AAC64368]99 99Gibberella zeae [AAV66343] Aspergillus fumigates [XP747393] Aspergillus clavatus [XP001276684]FungiAspergillus nidulans [XP660146]0.Figure 8 Unrooted phylogenetic tree of selected polysaccharide lyase family 3 proteins generated using maximum likelihood analysis Unrooted phylogenetic tree of selected polysaccharide lyase family 3 proteins generated using maximum likelihood analysis. The PL3 proteins from A. avenae (AA-PEL-1 and AA-PEL-2) are labeled Staurosporine in bold. GenBank accession numbers of PL3 proteins from Bursaphelenchus mucronatus (BM-PEL-1 and BM-PEL-2), Bursaphelenchus xylophilus (BX-PEL-1 and BX-PEL2), Meloidogyne incognita (MI-PEL-1, MI-PEL-2 and MI-PEL-3), Meloidogyne javanica (MJ-PEL), Globodera rostochiensis (GR-PEL-1 and GR-PEL-2), Heterodera schachtii (HS-PEL-1 and HS-PEL-2), bacteria and fungi are indicated in brackets. The bootstrap values are calculated from 1000 replicates. The scale bar represents 20 substitutions per 100 amino acid positions.Comparative analyses were performed with cluster sequences as queries versus multiple databases including C. elegans Wormpep v.203 protein database [46]. Protein databases for 'other nematode' and 'non-nematodes' were generated in-house for similarity searches. The 'other nematode'database contained all available protein sequences from nematodes other than C. elegans and A. avenae as well as nematode ESTs in GenBank (April 30, 2009), translated into peptide sequences (in TBLASTX analysis). The 'non-nematode' database comprises amino acid sequences from the complete non-redundant protein database (April 30, 2009) excluding those from nematodes. Homologues to cluster sequences were identifiedvia comparisons against WormBase and non-redundant protein databases using the BLASTX algorithm. The TBLASTX algorithm with default parameters was used to compare the A. avenae cluster sequences to ESTs from other nematodes. Each cluster sequence of A. avenae was assigned a 'statistically significant' match if the E-value from the BLAST output of the sequence alignment was PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/16989806 A. avenae genomic DNA, using pairs of gene-specific primers flanking each open reading frame. PCR products were cloned into the pGEM-T Easy vector (Promega) and sequenced using standard protocols. Signal peptide predictions were made using the SignalP program [63]. Protein theoretical pI and molecular mass were predicted using the "Compute pI/MW tool" available at ExPASy http://br.expasy.org/tools/pi_tool.html. Sequence alignments were made with a multiple alignment program, MAFFT version 6 http://align.bmr.kyushuu.ac.jp/mafft/software/about.html and output was produced using BOXSHADE 3.21 http://align.bmr.kyushuu.ac.jp/mafft/software/about.htmlPhylogenetic analyses The phylogenetic analyses of the catalytic domains of the GHF5 proteins and PL3 proteins were performed on the Phylogeny.fr platform [74] and comprised the following steps. Sequences were aligned.