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Cu 41l4

Manufactured by Percival
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Sourced in Germany, United States

The CU-41L4 is a laboratory equipment designed for specialized tasks. It features advanced capabilities to support various research and analytical applications. The core function of the CU-41L4 is to provide precise and reliable performance for users in controlled laboratory environments.

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Lab products found in correlation

Plant agar, by Duchefa Biochemie (3 mentions) Glass bead, by Merck Group (2 mentions) Tissuelyzer, by Qiagen (2 mentions) Sucrose, by Duchefa Biochemie (2 mentions) Tissue culture plates, by Techno Plastic Products (2 mentions) Sungro professional growing mix 1, by Sun Gro Horticulture (2 mentions) Murashige and skoog ms medium, by Duchefa Biochemie (2 mentions) Triton x 100, by Merck Group (1 mentions)

13 protocols using cu 41l4

1

Arabidopsis Bacterial Infection Assay

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A. thaliana seeds were sterilized using 70% ethanol for 1 min and stratified for 4 days at 4°C in the dark. Plants were grown in potting soil (Substrat 1, Klasmann-Deilmann, Germany) for 4-5 weeks in growth chambers (CU-41L4, Percival) fitted with full spectrum lights (Philips Master TL-D, 18 W/840) at 22°C and 60% relative humidity under an 11-hour light cycle. Bacteria were grown on R2A-MeOH agar plates for four days, restreaked and incubated for another three days at 22°C. Cells were resuspended in 10 mM MgCl2, and the OD600 was adjusted to 0.002 (~106 CFU/ml). Three to four leaves of four plants were infiltrated per treatment. Plants were covered with a plastic dome to increase humidity, and infection was monitored for 5 days.
Samples were taken from infiltrated leaves at the indicated time points. Leaves were surface sterilized by submerging them in 70% ethanol for 30 sec and subsequently washed in water three times. Leaf discs were taken with a cork borer (diameter 7.5 mm), and two leaf discs from the same plant were combined in tubes filled with 200 μl of 10 mM MgCl2 and two glass beads (3 mm diameter, Merck, Darmstadt, Germany). Leaf discs were homogenized with TissueLyzer (QIAGEN, Hilden, Germany) at 30 Hz for 2x 45 sec. The bacterial solution was plated as a dilution series on R2A-MeOH agar plates to count CFUs.
Pfeilmeier S., Petti G.C., Bortfeld-Miller M., Daniel B., Field C.M., Sunagawa S, & Vorholt J.A. (2021). The plant NADPH oxidase RBOHD is required for microbiota homeostasis in leaves. Nature microbiology, 6(7), 852-864.
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2

Arabidopsis Growth and Infection Protocol

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Arabidopsis thaliana Columbia (Col-0) and
bak1-5/bkk1-166 (link) were cultivated as described before31 (link). Seeds were
surface-sterilized, stratified in water at 4°C for 3-4 days before placed
in 24-well plates containing 1.5 ml of MS including vitamins (Duchefa)
supplemented with 3% w/v sucrose and 0.55% w/v plant agar (Duchefa). Plates were
sealed with parafilm and incubated in a growth chamber (Percival, CU41-L4) set
to 24°C/22°C and 65% relative humidity under long-day conditions
(16 h light/8 h dark) for one week prior to switching to short-day conditions (9
h light/15 h dark). Plates were shuffled 2-3 times a week. Parafilm was removed
one day before infection.
Vogel C.M., Potthoff D.B., Schäfer M., Barandun N, & Vorholt J.A. (2021). Protective role of the Arabidopsis leaf microbiota against a bacterial pathogen. Nature microbiology, 6(12), 1537-1548.
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3

Arabidopsis Growth and Infection Protocol

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Arabidopsis thaliana Columbia (Col-0) and
bak1-5/bkk1-166 (link) were cultivated as described before31 (link). Seeds were
surface-sterilized, stratified in water at 4°C for 3-4 days before placed
in 24-well plates containing 1.5 ml of MS including vitamins (Duchefa)
supplemented with 3% w/v sucrose and 0.55% w/v plant agar (Duchefa). Plates were
sealed with parafilm and incubated in a growth chamber (Percival, CU41-L4) set
to 24°C/22°C and 65% relative humidity under long-day conditions
(16 h light/8 h dark) for one week prior to switching to short-day conditions (9
h light/15 h dark). Plates were shuffled 2-3 times a week. Parafilm was removed
one day before infection.
Vogel C.M., Potthoff D.B., Schäfer M., Barandun N, & Vorholt J.A. (2021). Protective role of the Arabidopsis leaf microbiota against a bacterial pathogen. Nature microbiology, 6(12), 1537-1548.
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4

Cultivation of Arabidopsis thaliana in Controlled Conditions

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Plants were cultivated as described previously37 (link), with some modifications. In brief,
Arabidopsis thaliana Columbia-0 (Col-0) seeds were
surface-sterilized67 and stratified for 4 days at 4°C. Plants were cultivated in six- or
twelve-well tissue culture plates (TechnoPlasticProducts) filled with 5 or 2 ml
washed and heat-sterilized calcined clay and 2.5 or 1 ml half-strength Murashige
& Skoog medium pH 5.8 including vitamins (½ MS, Duchefa),
respectively. Individual seeds were placed in the center of each well. If seeds
did not germinate, a seedling was transplanted to the corresponding well six
days after seeding. Starting at day 6, each well was supplemented twice per week
with 200 or 100 μL ½ MS, respectively. Plates were incubated in a
growth chamber (Percival, CU41-L4) set to 22°C and 54% relative humidity
with a 11 h light 13 hours dark regime fitted with full spectrum lights (Philips
Master TL-D 18W/950 Graphica) and lights emitting a higher fraction of UVA and
UVB (Sylvania Reptistar F18W/6500K). Combined light intensity was set to 200-210
µmol m-2 s-1 for wavelength 400-700 nm and 4-5
µmol m-2 s-1 for wavelength 280-400 nm.
Schäfer M., Vogel C.M., Bortfeld-Miller M., Mittelviefhaus M, & Vorholt J.A. (2022). Mapping phyllosphere microbiota interactions in planta to establish genotype-phenotype relationships. Nature microbiology, 7(6), 856-867.
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5

Arabidopsis Leaf Degradation Assay

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A. thaliana wild-type Col-0, bbc30 (link), fls2/efr/cerk131 (link), rbohD knockout mutant17 (link) and complementation line rbohD/pRBOHD::RBOHD-FLAG (rbohD/RBOHD)65 (link) were used in this study.
A. thaliana plants for leaf degradation assays were grown in peat-based potting soil (substrate 1, Klasmann-Deilmann) in a growth chamber (CU-41L4, Percival) under controlled conditions (11 h light cycle, 22 °C, 65% relative humidity, light intensity (photosynthetic active radiation) 200 µmol s−1 cm−2). Seeds were treated with 70% ethanol for 2 min, sown on soil and stratified for 2 days at 4 °C in the dark.
Pfeilmeier S., Werz A., Ote M., Bortfeld-Miller M., Kirner P., Keppler A., Hemmerle L., Gäbelein C.G., Petti G.C., Wolf S., Pestalozzi C.M, & Vorholt J.A. (2024). Leaf microbiome dysbiosis triggered by T2SS-dependent enzyme secretion from opportunistic Xanthomonas pathogens. Nature Microbiology, 9(1), 136-149.
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6

Characterizing Arabidopsis Homozygous Mutants

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Homozygous kea1-1 kea2-1 (CS72318) and kea1-2 kea2-2 (CS72319) were obtained from our previous study (Kunz et al., 2014 (link)). Except for rps5-2, all other loss-of-function lines were obtained from ABRC. Lines were confirmed to be homozygous using PCR with one gene-specific primer and one T-DNA-specific primer. For a full list of lines and primers, see Supplemental Table S4. Seeds were sterilized in 70% (v/v) ethanol, plated on ½ concentration Murashige and Skoog (MS) media with 0.8% (w/v) agar and stratified in the dark at 4°C for 48 h (Murashige and Skoog, 1962 ). Plates were then placed in growth chamber (CU-41L4; Percival Scientific) with 150 µmol photons m−2s−1 of continuous light (cool-white fluorescent bulbs), 16-h:8-h light: dark cycle. Growth temperatures were 22°C in the light and 18°C in the dark. Plants were grown for 7 days and then either transferred to soil (Sungro Professional Growing Mix #1, Sun Gro Horticulture, USA) or to ½ MS agar plates with additional treatment. Plants transferred to soil were grown at the same light intensity, light cycle, and temperature described above in a Bio Chamber SPC-56 with florescent tubes (Phillips FS4T5/841HO) and LED bulbs (Satco LED A19 15.5 watt, 2700K). Treatment plates contained 67.5mM NaCl or respective NaCl or KCl amounts as given in the figure legend.
DeTar R.A., Barahimipour R., Manavski N., Schwenkert S., Höhner R., Bölter B., Inaba T., Meurer J., Zoschke R, & Kunz H.H. (2021). Loss of inner-envelope K+/H+ exchangers impairs plastid rRNA maturation and gene expression. The Plant Cell, 33(7), 2479-2505.
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7

Arabidopsis Bacterial Infection Assay

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A. thaliana seeds were sterilized using 70% ethanol for 1 min and stratified for 4 days at 4°C in the dark. Plants were grown in potting soil (Substrat 1, Klasmann-Deilmann, Germany) for 4-5 weeks in growth chambers (CU-41L4, Percival) fitted with full spectrum lights (Philips Master TL-D, 18 W/840) at 22°C and 60% relative humidity under an 11-hour light cycle. Bacteria were grown on R2A-MeOH agar plates for four days, restreaked and incubated for another three days at 22°C. Cells were resuspended in 10 mM MgCl2, and the OD600 was adjusted to 0.002 (~106 CFU/ml). Three to four leaves of four plants were infiltrated per treatment. Plants were covered with a plastic dome to increase humidity, and infection was monitored for 5 days.
Samples were taken from infiltrated leaves at the indicated time points. Leaves were surface sterilized by submerging them in 70% ethanol for 30 sec and subsequently washed in water three times. Leaf discs were taken with a cork borer (diameter 7.5 mm), and two leaf discs from the same plant were combined in tubes filled with 200 μl of 10 mM MgCl2 and two glass beads (3 mm diameter, Merck, Darmstadt, Germany). Leaf discs were homogenized with TissueLyzer (QIAGEN, Hilden, Germany) at 30 Hz for 2x 45 sec. The bacterial solution was plated as a dilution series on R2A-MeOH agar plates to count CFUs.
Pfeilmeier S., Petti G.C., Bortfeld-Miller M., Daniel B., Field C.M., Sunagawa S, & Vorholt J.A. (2021). The plant NADPH oxidase RBOHD is required for microbiota homeostasis in leaves. Nature microbiology, 6(7), 852-864.
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8

Arabidopsis Genotypes Under Light Conditions

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A. thaliana WT and mutant plants (accession Columbia‐0 [Col‐0]) were germinated on one‐half Murashige and Skoog (MS), 1% phytoagar plates pH 5.8 for 7 days in a growth chamber (CU‐41L4; Percival Scientific) with 150 μmol photons m−2 s−1 of continuous light (cool‐white fluorescent bulbs), 16/8‐h light:dark cycle. After potting plants on soil (Sungro Professional Growing Mix #1, Sun Gro Horticulture, Agawam, MA, USA), they were shifted to self‐built growth racks with 120 μmol photons m−2 s−1 of continuous light in 12/12‐h light:dark cycles and room temperature 22°C (Schneider et al., 2019 (link)). When individuals were designated for light treatments, 2‐week‐old plants were moved either into constant high light (900 μmol photons m−2 s−1) or into fluctuating light for 2 weeks. The SALK_008491 (nhd1‐1) and Sail107_F07 aka nhd1‐2 were originally obtained from ABRC but used in an earlier study (Muller et al., 2014 (link)). The nhd1‐1nhd1‐2 mutant line was generated by crossing of respective single mutants. Subsequent BASTA selection of progenies ensured that both insertions were maintained in a heterozygous state. As described previously, this results in an another viable overall homozygous nhd1 loss‐of‐function allele (Muller et al., 2014 (link)).
Lopez L.S., Völkner C., Day P.M., Lewis C.M., Lewis C.L., Schneider D., Correa Galvis V., Cruz J.A., Armbruster U., Kramer D.M, & Kunz H. (2022). The Arabidopsis T‐DNA mutant SALK_008491 carries a 14‐kb deletion on chromosome 3 that provides rare insights into the plant response to dynamic light stress. Plant Direct, 6(7), e429.
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9

Cultivation of Arabidopsis thaliana in Controlled Conditions

Cited 1 time
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Plants were cultivated as described previously37 (link), with some modifications. In brief,
Arabidopsis thaliana Columbia-0 (Col-0) seeds were
surface-sterilized67 and stratified for 4 days at 4°C. Plants were cultivated in six- or
twelve-well tissue culture plates (TechnoPlasticProducts) filled with 5 or 2 ml
washed and heat-sterilized calcined clay and 2.5 or 1 ml half-strength Murashige
& Skoog medium pH 5.8 including vitamins (½ MS, Duchefa),
respectively. Individual seeds were placed in the center of each well. If seeds
did not germinate, a seedling was transplanted to the corresponding well six
days after seeding. Starting at day 6, each well was supplemented twice per week
with 200 or 100 μL ½ MS, respectively. Plates were incubated in a
growth chamber (Percival, CU41-L4) set to 22°C and 54% relative humidity
with a 11 h light 13 hours dark regime fitted with full spectrum lights (Philips
Master TL-D 18W/950 Graphica) and lights emitting a higher fraction of UVA and
UVB (Sylvania Reptistar F18W/6500K). Combined light intensity was set to 200-210
µmol m-2 s-1 for wavelength 400-700 nm and 4-5
µmol m-2 s-1 for wavelength 280-400 nm.
Schäfer M., Vogel C.M., Bortfeld-Miller M., Mittelviefhaus M, & Vorholt J.A. (2022). Mapping phyllosphere microbiota interactions in planta to establish genotype-phenotype relationships. Nature microbiology, 7(6), 856-867.
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10

Gnotobiotic Arabidopsis Growth and Inoculation

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A. thaliana ecotype Col‐0 seeds were surface sterilized and stratified at 4°C in the dark for 4 days (Innerebner et al., 2011). Sterile seeds were sowed in microboxes (O118/80+OD118 filter: #40, Combiness, Nazareth, Belgium) containing calcined clay (Diamond Pro Calcined Clay Drying Agent, Arlington, USA) as described previously (Bai et al., 2015). Plants were grown under gnotobiotic conditions at 22°C, 11 h light and 54% humidity in standard plant growth chambers (CU‐41L4, Percival, Perry, USA). After two weeks of growth, plants were inoculated with M. extorquens PA1. For inoculation, bacteria were grown on minimal medium plates containing 1.5% agar, harvested and solubilized in 10 mM MgCl2 and adjusted to OD600 0.007. Each plant was inoculated with 1 ml of bacteria solution or mock treated with 10 mM MgCl2. Plants were harvested after 28 d of growth and the above‐ground parts were carefully separated from roots using sterilized razor blades. For further processing, refer to the element analysis and proteomics parts.
Ochsner A.M., Hemmerle L., Vonderach T., Nüssli R., Bortfeld‐Miller M., Hattendorf B, & Vorholt J.A. (2019). Use of rare‐earth elements in the phyllosphere colonizer Methylobacterium extorquens PA1. Molecular Microbiology, 111(5), 1152-1166.
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