diff --git a/assays/Isolation of extracellular polysaccharide matrix/dataset/SuppFigS1.png b/assays/Isolation of extracellular polysaccharide matrix/dataset/SuppFigS1.png new file mode 100644 index 0000000000000000000000000000000000000000..734f797f47b9cc69f5dfa1bab12493f4b7aee1ff --- /dev/null +++ b/assays/Isolation of extracellular polysaccharide matrix/dataset/SuppFigS1.png @@ -0,0 +1,3 @@ +version https://git-lfs.github.com/spec/v1 +oid sha256:02a603268108fbf90e8df56d94abd5f991e128f27c54bf94818ce1d5dc5c1401 +size 208207 diff --git a/assays/Isolation of extracellular polysaccharide matrix/isa.assay.xlsx b/assays/Isolation of extracellular polysaccharide matrix/isa.assay.xlsx index 9ae2735534d88df36625cefa40e389c7a10f2c8c..245265e1af2ac5d00dac4846571b45a3345c195a 100644 Binary files a/assays/Isolation of extracellular polysaccharide matrix/isa.assay.xlsx and b/assays/Isolation of extracellular polysaccharide matrix/isa.assay.xlsx differ diff --git a/assays/Isolation of extracellular polysaccharide matrix/protocols/Isolation of extracellular polysaccharide matrix.md b/assays/Isolation of extracellular polysaccharide matrix/protocols/Isolation of extracellular polysaccharide matrix.md new file mode 100644 index 0000000000000000000000000000000000000000..64c7e5de71e899550b83b97ef6d8ac299fd0a939 --- /dev/null +++ b/assays/Isolation of extracellular polysaccharide matrix/protocols/Isolation of extracellular polysaccharide matrix.md @@ -0,0 +1,45 @@ +## Microbial strains and culture conditions for EPS matrix production in S. indica and B. sorokiniana + +The EPS matrix was isolated from the S. indica strain +expressing GFP in the dikaryotic wild-type background +DSM11827 (Wawra et al., 2016) and from the B. sorokiniana +wild-type strain ND90r. Serendipita indica spores were iso- +lated from 3-week-old cultures grown on solid complex me- +dium (Si_CM) using 0.002% (v/v) Tween water (Zuccaro +et al., 2011). For the preculture, 2 mL of S. indica spores at a +concentration of 500,000 mL–1 were inoculated in 100 mL +of TSB medium containing 1% (w/v) sucrose and shaking at +120 rpm at 28 C. After 48 h, the pre-cultures were trans- +ferred to 400 mL of TSB containing 1% sucrose and shaken +at 120 rpm at 28 Cfor 72 h. Bipolaris sorokiniana spores +were isolated from 10-day-old cultures grown on solid com- +plete medium (Bs_CM), using 0.002% (v/v) Tween water +(Sarkar et al., 2019). The spores were inoculated at a final +concentration of 250 spores mL–1 in 250 mL of YPD me- +dium and these samples were shaken at 28 C for 36 h. + +## Isolation of EPS matrix from S. indica and B. sorokiniana culture supernatant + +Culture supernatants from axenically grown S. indica or B. +sorokiniana were collected by filtering the mycelia using +Miracloth (Merck Millipore). The EPS matrix was isolated +from the culture media using cryogelation. Briefly, the cul- +ture media were frozen overnight at –20°C and slowly +thawed at room temperature for 16 h. The precipitated EPS matrix (Supplemental Figure S1) present in the culture me- +dium was isolated using a pipette controller and washed +four times with 30 mL of Milli-Q water and either used for +proteome analyses (see section “Proteome analysis of +S.indica EPS matrix, CW, and culture filtrateâ€) or washed +one more time with 30 mL of 8.3-mM EDTA (pH 8.0) to re- +move metal ions potentially present in the EPS. The proteins +present in the EPS matrix were removed by treatment with +30 mL of protein denaturation solution (containing 8-M +urea, 2-M thiourea, 4% [w/v] sodium dodecyl sulfate [SDS], +100-mM Tris–HCl, pH 7.5) and boiling at 95°C for 15 min. +The SDS present in the EPS matrix was removed by boiling +the material with 30 mL of Milli-Q water at 95°C for 10 min +and centrifuging at 10,000g for 10 min at room temperature. +The latter step was repeated approximately 15 times until +no further foaming occurred. The resulting protein-free EPS +matrix was lyophilized and used for glycosyl linkage, TLC or +MALDI-TOF analyses. \ No newline at end of file diff --git a/assays/Proteome analysis of S indica EPS matrix CW and cullture filtrate/protocols/Proteome analysis of S. indica EPS matrix, CW, and culture filtrate b/assays/Proteome analysis of S indica EPS matrix CW and cullture filtrate/protocols/Proteome analysis of S. indica EPS matrix, CW, and culture filtrate new file mode 100644 index 0000000000000000000000000000000000000000..e69de29bb2d1d6434b8b29ae775ad8c2e48c5391 diff --git a/studies/Characterization of fungal components/isa.study.xlsx b/studies/Characterization of fungal components/isa.study.xlsx index f85cffafe62cce8afd6d068e39d2150aad0bed59..c65e3723e1d81b370189cb75e511861c162c3e1f 100644 Binary files a/studies/Characterization of fungal components/isa.study.xlsx and b/studies/Characterization of fungal components/isa.study.xlsx differ