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Bck1 and their orthologous, and the MEGA version 5.2 was used to produce the phylogenetic tree. Bootstrap values were expressed as a percentage of 1000 replicates. ORF and exon/intron positions were identified by comparing genomic and cDNA sequences. Functional domains of FoSlt2, FoMkk2 and FoBck1 genes were determined using Conserved Domain Architecture Retrieval Tool available from NCBI database. Fusaric acid quantification For liquid medium fermentation, 107 microconidia of WT and mutants FoSlt2, FoMkk2, and FoBck1 were inoculated in 250 mL Czapek Dox medium and incubated at 28C on a rotary shaker at 170 rpm for 9 days, the cultures were filtrated with Waterman filters to exclude mycelium and conidia, and crude mycotoxion was extracted as described. An Agilent 1260 RP-HPLC system with an Agilent HC-C18 column was employed to analysis fusaric acid. Elution was carried out using a mobile phase KU-55933 site comprising 20% methanol, 48% double distilled water and 32% H3PO4 for 40 min with a UV detector at 280 nm, the flow rate was 1 mL/min. Before injection, the samples were filtrated through 0.45-m filters. For solid medium fermentation, 250 mL glass bottles containing 50 g of grains mixture and 25mL of distilled water were stopped with cotton plugs and autoclaved at 121C for 30 min, glass bottles were inoculated with 108 microconidia of WT and the three mutants and cultured at 28C, respectively. After 23 days, the grains cultures were collected and dried. The crude mycotoxion was extracted from 2 g grains cultures with 10 mL acetonitrile/ water/acetic acid by sonication for 20 min, and then supernatant and residue were separated. Fusaric acid was analysed as above described with some modifications, 60% methanol, 5% double distilled water and 35% H3PO4 was used for mobile phase. 18 / 24 Roles of MAP Kinases in F. oxysporum f. sp. cubense Virulence assay The virulence of the three gene deletion PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19768747 mutants was tested on tissue culture-derived banana plantlets at the 45 leaf stage. Banana root inoculation assays were performed as described. Disease symptoms were assessed 40 days after inoculation. Thirty plantlets were used for each treatment. Data analysis Statistical analysis of the data was carried out using SPSS 21.0 software, with a completely randomized analysis of the variances. Tukey’s honest significant difference test was used for comparison of the means. Accession Numbers Sequence data can be found in the Fusarium Comparative Genome database under the following accession numbers: FoSlt2, FOIG_09199; FoMkk2, FOIG_05686; FoBck1, FOIG_03241; actin, FOIG_02823; seven chitin synthase genes, FOIG_07229, FOIG_10825, FOIG_09216, FOIG_00580, FOIG_06735, FOIG_06738 and FOIG_06723; four peroxidase synthase genes, FOIG_08821, FOIG_07465, FOIG_04532 and FOIG_09161; beas, FOIG_15793; kivr, FOIG_15792; abc3, FOIG_15791; sidA, FOIG_11772; FUB1, FOIG_16450; FUB2, FOIG_16451; FUB3, FOIG_16452; FUB4, FOIG_16453; FUB5, FOIG_16454. ~~ Astroviruses were first identified by Appleton and Higgins in 1975, in association with an outbreak of infantile gastroenteritis in a maternity ward in England and the Astroviridae family today contains viruses which infect more than 44 animal species. Human astroviruses are PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19768759 recognized as common viral pathogens causing gastroenteritis in infants and young children, with very few reports of HAstV-mediated disease in normal healthy adults, and some reports of severe disease after dissemination to extra-intestinal tissues in immunocompro

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Author: Cholesterol Absorption Inhibitors