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MB Sample ID: SA167616
Local Sample ID: | SAMP07_ES1 |
Subject ID: | SU001882 |
Subject Type: | Fungi |
Subject Species: | Exidia glandulosa;Mucidula mucida |
Taxonomy ID: | 5219;139077 |
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Subject:
Subject ID: | SU001882 |
Subject Type: | Fungi |
Subject Species: | Exidia glandulosa;Mucidula mucida |
Taxonomy ID: | 5219;139077 |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
SAMP07_ES1 | SA167616 | FL019928 | Stable Temp | Treatment |
Collection:
Collection ID: | CO001875 |
Collection Summary: | The two fungal strains M. mucida and E. glandulosa were isolated from dead branches attached to standing beech (F. sylvatica) trees at Clyne Valley Woodlands, a minimally managed woodland in Swansea, South Wales, UK (Lat 51.6063, Long -4.0068). Isolates were collected from different but closely positioned trees within the same woodland stand. |
Sample Type: | new |
Collection Method: | Woodchip isolation |
Collection Location: | Clyne Valley Woodlands, Swansea, United Kingdom, Lat 51.6063, Long -4.0068 |
Treatment:
Treatment ID: | TR001895 |
Treatment Summary: | F. sylvatica blocks (John Harrison, Wrexham, UK) with dimensions 2 cm3 were autoclaved three times and then pre-colonised by placing them on 0.5% malt-extract agar (MEA; 0.5% malt, 1.5% agar w/v; Sigma-Aldrich, Dorset, UK) cultures of the appropriate fungal strain and incubated in the dark at 20°C. Following 8-weeks pre-colonisation, experimental treatments consisting of 2 x 2 block matrices colonised with one of the two fungal strains per microcosm. Microcosms were subjected to a diurnally cycling temperature sequence of 5, 15, 25 and 15°C per 24 hours so that temperature changed by 10°C every 6 hours with a mean of 15 ± 10°C. Temperatures were chosen to represent the minima and maxima canopy-dwelling ligninolytic fungi encounter during the growing season in a temperate woodland ecosystem. Control microcosms were kept at a constant temperature of 15°C for the duration of the experiment. These methods were also chosen to be comparable with those of Toljander Lindahl, Holmer & Högberg (Toljander et al., 2006). All microcosms were incubated in the dark for 8 weeks with three replicates per species per treatment (n = 12). |
Sample Preparation:
Sampleprep ID: | SP001888 |
Sampleprep Summary: | At the end of the experiment, each block was split into three segments from top to bottom using a sterile chisel. The top and middle sections were quenched in liquid nitrogen, lyophilised and stored at -20°C. From the bottom third of each block, four wood chips (~0.5 mm2) were extracted, one from each quadrant, and placed onto 2% (w/v) MEA. Cultures were incubated at 20°C for 7 – 10 days and mycelial morphology confirmed that no woodblocks had become contaminated during the experiment. One third of the frozen woodblock sections from each treatment group were each weighed and manually chipped, added to ≤ 10 ml acetonitrile : methanol : H2O (2:2:1) and shaken at 180 rpm at room temperature for 1 h. Extracts were filtered through glass wool and Whatman filter paper (no. 1) and centrifuged at 3,500 g for 30 min. The supernatant was dried in vacuo (Eppendorf, Hamburg, Germany) overnight. Dried samples were derivitized by addition of 30 μl methoxylamine hydrochloride (15 mg ml-1 in pyridine; Sigma, UK) followed by incubation at 60°C for 90 min. Subsequently 50 μl MSTFA+ 1% (v/v) TMCS (Thermo Scientific) was added to the sample and incubated at 40°C for 60 min. Derivatized samples were transferred to autosampler vials and 10 μl tetracosane (5 mg ml-1 in heptane; Sigma-Aldrich) added as an internal standard. |
Combined analysis:
Analysis ID | AN002927 |
---|---|
Analysis type | MS |
Chromatography type | GC |
Chromatography system | Agilent 5975 |
Column | Phenomenex Zebron ZB-WAXplus (30m x 0.25mm,0.25um) |
MS Type | EI |
MS instrument type | Single quadrupole |
MS instrument name | Agilent 5975 |
Ion Mode | POSITIVE |
Units | Integrated signal (SpectConnect) |
Chromatography:
Chromatography ID: | CH002169 |
Instrument Name: | Agilent 5975 |
Column Name: | Phenomenex Zebron ZB-WAXplus (30m x 0.25mm,0.25um) |
Column Temperature: | 250 |
Flow Rate: | 1ml/min |
Internal Standard: | Tetracosane |
Chromatography Type: | GC |
MS:
MS ID: | MS002718 |
Analysis ID: | AN002927 |
Instrument Name: | Agilent 5975 |
Instrument Type: | Single quadrupole |
MS Type: | EI |
MS Comments: | - |
Ion Mode: | POSITIVE |