V. unguiculata (Burpee black-eyed pea no. 5 plants) were seeded in 3–1/4” square pots (Greenhouse Megastore) using Pro Mix BX soil (Greenhouse Megastore) and maintained in Conviron A1000 Reach-in growth chambers (Conviron) at 16/8 hour day/night cycles at 25 °C/22 °C and 60% relative humidity. When the plants were 10 days old, the primary leaves were mechanically inoculated with 40 μL (0.1 mg mL−1) of native or inactivated CPMV (see above); the leaves were lightly dusted with carborundum (Thermo Fisher Scientific) to promote mechanical lesions on the leaves enabling the virus infection. 10 plants were used per treatment arm; in addition, 10 plants were not treated. Leaves were imaged and harvested 10 days post-inoculation and stored at −80 °C until further processing.
Carborundum
Manufactured by Thermo Fisher Scientific
Sourced in United States
Carborundum is a synthetic crystalline material composed of silicon and carbon. It is an abrasive and refractory material known for its hardness and thermal resistance. Carborundum is commonly used in the production of various industrial and laboratory equipment.
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Lab products found in correlation
8 protocols using carborundum
1
Pea Virus Infection Assay
2
Mechanical Inoculation of Tobacco with Plant Viruses
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N. tabacum cv. Samsun NN plants were germinated and cultivated in the growth room maintained with a 16 h light/8 h dark cycle, a daytime temperature of 24°C, and a nighttime temperature of 22°C. The plants at the 5 to 6 leaf stage were inoculated mechanically with sap inoculum of TMV-U1, Fny-CMV, potato virus X (PVX-Kr), or potato virus Y (PVY-O) (viruses as described in Baek et al., 2017 (link)), after dusting with Carborundum (600 mesh, Thermo Fisher Scientific, Waltham, MA, USA) using 10 mM phosphate buffer, pH 7.2.
3
Pea Virus Inoculation Protocol
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V. unguiculata (Burpee black-eyed pea no. 5 plants) were seeded in 3–1/4′′ square pots (Greenhouse Megastore) using Pro Mix BX soil (Greenhouse Megastore) and maintained in Conviron A1000 Reach-in growth chambers (Conviron) at 16/8 hour day/night cycles at 25 °C/22 °C and 60% relative humidity. When the plants were 10 days old, the primary leaves were mechanically inoculated with 40 μL (0.1 mg mL−1) of native or inactivated CPMV (see above); the leaves were lightly dusted with carborundum (Thermo Fisher Scientific) to promote mechanical lesions on the leaves enabling the virus infection. 10 plants were used per treatment arm; in addition, 10 plants were not treated. Leaves were imaged and harvested 10 days post-inoculation and stored at −80 °C until further processing.
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V. unguiculata (Burpee black-eyed pea no. 5 plants) were seeded in 3–1/4′′ square pots (Greenhouse Megastore) using Pro Mix BX soil (Greenhouse Megastore) and maintained in Conviron A1000 Reach-in growth chambers (Conviron) at 16/8 hour day/night cycles at 25 °C/22 °C and 60% relative humidity. When the plants were 10 days old, the primary leaves were mechanically inoculated with 40 μL (0.1 mg mL−1) of native or inactivated CPMV (see above); the leaves were lightly dusted with carborundum (Thermo Fisher Scientific) to promote mechanical lesions on the leaves enabling the virus infection. 10 plants were used per treatment arm; in addition, 10 plants were not treated. Leaves were imaged and harvested 10 days post-inoculation and stored at −80 °C until further processing.
4
Detecting Viral Infection in Cowpea Leaves
5
Chili Pepper Inoculation with CMV
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CMV-GTN was maintained in Nicotiana tabacum leaves and confirmed virus viability by the immunostrip method (Choi et al., 2015 ). Infected leaves were collected aseptically and ground with 0.01 M phosphate buffer (pH 7.0) using a sterilized plastic pouch in the ratio of 1 g leaf tissue per 10 ml of the buffer. The suspension is used as the source of inoculum for chili pepper infection. The first two true leaves of each chili pepper plant were lightly dusted with carborundum (Thermo Fisher Scientific, Waltham, MA, USA) and then rubbed with inoculum from leaf base to tip, contains CMV-infected tobacco leaf suspension. All the plants including the controls were inoculated with CMV.
6
Melon Virus Susceptibility and RDR Gene Mapping
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Cucumis melo L. cvs. Arava and Vedrantais were used for CmRDR gene mapping and virus susceptibility analyses. In addition, in 10 melon genotypes (listed in Table S1 ) the truncated RDR1b genes were mapped. Seeds were planted in a soil mixture and grown in a greenhouse in daylight at 25 °C. The viruses listed in Table S2 were used for virus accumulation analysis and evaluation of RDR gene expression, namely: cucumber mosaic virus CMV—Fny strain, CMV mutant lacking the 2b gene (CMV-Δ2b) [36 (link)], CGMMV, CVYV, ZYMV, PRSV-W and WMV. Squash (Cucurbita pepo L. cv. Zuccini) plants were a source of inocula for PRSV-W, WMV and ZYMV. Cucumber (Cucumis sativus L. cv. Bet Alfa) plants were inocula sources for CMV, CMV-Δ2b, CGMMV and CVYV. Melon seedlings at the cotyledon stage with a small true leaf (about 3–5 days post-emergence) were dusted with carborundum (320 grit powder, Fisher Scientific, USA) before mechanical inoculation with virus-bearing sap (ca. 1:10 ratio g tissue/mL H2O) [37 (link)].
7
Pea Virus Inoculation Procedure
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Seeds of P. sativum (organic pea Progress #9, Garden Harvest Supply, Berne, IN, USA) were sown individually in 5.0 × 5.5 × 7 cm pots with Sunshine LC-1 mix soil (Professional growing mix—73–83% Canadian sphagnum peat moss, perlite, dolomite lime, SHU 521) in a climate chamber (20 ± 2 °C, 12 h light at approximatively 60 µmol·m−2·s−1). Seven days after sowing, seedlings were mechanically inoculated with 5 µL of two equal amounts of either PEMV1 or CP mutant and PEMV2 in vitro transcripts (IVT) using 800 ng of each IVT. IVTs mixes were rubbed on the first leaves using an abrasive powder (Carborundum, Fisher Scientific Co., Pittsburgh, PA, USA). The inoculated and newly emerged leaves of each plant were sampled for measurement of viral accumulation. The positions of all inoculated plants for the experiment were randomized in the climate chamber. Control plants were mock-inoculated with 5 µL of RNA-free water.
8
Tobacco Leaf Virus Inoculation Protocol
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Freshly infected leaves from tobacco plants were pre-frosted in liquid nitrogen and ground [tissue: buffer; 1:6 (weight/ volume)] with cold and freshly prepared potassium phosphate buffer (0.01 M), having pH of 7.0, consisting of 0.01 M 2-mercaptoethanol and 0.2% sodium sulfite, in a pre-chilled pestle and mortar. The ground inoculum was filtered through a muslin cloth to remove the debris. Celite 545 (Fisher Scientific) and Carborundum (Fisher Scientific) with a ratio of 1:2. The prepared inoculum was placed on ice till its use for inoculation of the test plants [52 (link)].
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