Bioinformatic analysis of MLO gene family in rose

The chromosomal location and predicted intron/exon structure of each RhMLO gene were extracted from the available genomic information of Rosa chinensis genome HapOB2 v1.0 in GDR. We visualized the chromosomal location and intron/exon structure by MapChart 2.32 and online tool GSDS 2.0 (http://gsds.cbi.pku.edu.cn/), respectively.

A phylogenetic analysis was performed for the RhMLO proteins obtained from the BLAST search. For this, next to the MLO sequences obtained from rose, apple, strawberry, peach and Arabidopsis, we also included a series of MLO homologs that have been functionally associated with powdery mildew susceptibility: HvMLO ({"type":"entrez-nucleotide","attrs":{"text":"Z83834","term_id":"1877220","term_text":"Z83834"}}Z83834), OsMLO2 ({"type":"entrez-nucleotide","attrs":{"text":"AF384030","term_id":"15290604","term_text":"AF384030"}}AF384030), TaMLO1_A1b ({"type":"entrez-nucleotide","attrs":{"text":"AX063298","term_id":"12541088","term_text":"AX063298"}}AX063298), TaMLO_B1 ({"type":"entrez-nucleotide","attrs":{"text":"AF361932","term_id":"15290588","term_text":"AF361932"}}AF361932), SlMLO1 ({"type":"entrez-protein","attrs":{"text":"AAX77013","term_id":"62208139","term_text":"AAX77013"}}AAX77013), CaMLO2 ({"type":"entrez-protein","attrs":{"text":"AFH68055","term_id":"384113234","term_text":"AFH68055"}}AFH68055), PsMLO1 ({"type":"entrez-nucleotide","attrs":{"text":"FJ463618","term_id":"225698199","term_text":"FJ463618"}}FJ463618), MtMLO1 ({"type":"entrez-nucleotide","attrs":{"text":"HQ446457","term_id":"318101906","term_text":"HQ446457"}}HQ446457), LjMLO1 ({"type":"entrez-protein","attrs":{"text":"AAX77015","term_id":"62208143","term_text":"AAX77015"}}AAX77015), VvMLO6 (Genoscope ID: GSVIVT00018217001; http://www.genoscope.cns.fr/externe/GenomeBrowser/Vitis/entry_ggb.html), VvMLO7 (Genoscope ID: GSVIVT00018219001), CsMLO1 (Cucurbit Genomics Database ID: Csa5M623470.1; http://cucurbitgenomics.org/organism/2) and NtMLO1 ({"type":"entrez-protein","attrs":{"text":"AIT98396","term_id":"698628402","term_text":"AIT98396"}}AIT98396) (Buschges et al. 1997; Elliott et al. 2002; Bai et al. 2008; Feechan et al. 2008; Winterhagen et al. 2008; Humphry et al. 2011; Zheng et al. 2013; Berg et al. 2015; Nie et al. 2015; Pessina et al. 2016b).

MLO protein sequences were aligned first with Clustalx 1.83 and then further aligned by Clustalw in MEGA7. Sequences were trimmed at the C and N terminal parts. Different amino acid substitution models were tested, and a Poisson model with uniform rates was chosen as most suitable. Then, the alignment was used to generate phylogenetic trees in MEGA7 using ML (maximum likelihood) and UPGMA (unweighted pair group method with arithmetic mean) methods with a bootstrap value of 1000.