Nightshade lb 985 plant imaging system

Manufactured by Berthold Technologies

Sourced in Germany

The NightShade LB 985 Plant Imaging System is a laboratory equipment designed for capturing high-quality images of plants. It features a specialized imaging chamber and software for acquiring and analyzing plant-related data.

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Lab products found in correlation

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15 protocols using nightshade lb 985 plant imaging system

Luminescence Imaging of Pea Seeds

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Pea seeds were imaged by employing the NightShade LB 985 Plant Imaging System (Berthold Technologies, Bad Wildbad, Germany) equipped with a highly sensitive CCD-sensor (NightOWLcam, Berthold Technologies) thermoelectrically-cooled to −74 °C to minimise thermal noise. The camera was mounted on a dark, light-tight chamber into which the seeds to be imaged were placed (approximately 20 cm away from the camera lens). Luminescence emissions from the control seeds and weevil-treated seeds were imaged and photons emanating from the samples were detected with a back-lit, midband-coated full frame chip possessing a spectral range of 350–1050 nm (with quantum efficiency of 90% at 620 nm wavelength). During image acquisition the binning factor was set to: 2 × 2 via the software, hence, the images were captured at the resolution of 512 × 512 pixels at a final 26 × 26 µm² pixel size (slow scan mode). The exposure time was set to 60 s. For image analysis the IndiGo software (V. 2.0.5.0, Berthold Technologies, Germany) was used. Before each measurement, all the samples were kept in the light-tight dark chamber for 1 h.
The CPS values of healthy and damaged samples caused by B. pisorum were examined statistically by one-way ANOVA. Means were separated by using the Tukey (HSD) test, at p ≤ 0.05.

Keszthelyi S., Fajtai D., Pónya Z., Somfalvi-Tóth K, & Donkó T. (2021). A Non-Invasive Approach in the Assessment of Stress Phenomena and Impairment Values in Pea Seeds Caused by Pea Weevil. Plants, 10(7), 1470.

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Firefly LCI Assay in Nicotiana benthamiana

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The firefly LCI assays were performed using N. benthamiana leaves. Plasmids for LCI were described previously (Liu et al., 2017 (link), 2019 (link)). Both the nLUC- (N-terminal luciferase) and cLUC- (C-terminal luciferase) fusion constructs with or without pSPYNE-FHY3 (empty vector pSPYNE as control) were co-infiltrated into N. benthamiana leaves via A. tumefaciens-mediated co-infiltration. The infiltrated plants were incubated for 2 or 3 days before examining using the Night SHADE LB 985 Plant Imaging System (Berthold, German).

Xie Y., Ma M., Liu Y., Wang B., Wei H., Kong D, & Wang H. (2021). Arabidopsis FHY3 and FAR1 Function in Age Gating of Leaf Senescence. Frontiers in Plant Science, 12, 770060.

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Ultra-weak Photon Emission Imaging of Cucumber Leaves

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Leaves of cucumbers grown under different nitrate supply levels were subjected to UPE (ultra-weak photon emission) imaging. Intact leaves of approximately the same size were separated from the plants and placed in the NightShade LB 985 Plant Imaging System (Berthold Technologies, Bad Wildbad, Germany). Luminescence emission of photons from the plants were visualized using a thermoelectrically cooled (−70 °C) CCD camera (NightOWLcam, Berthold Technologies) mounted on a dark, light-proof chamber. A back-lit, midband-coated full-frame chip with a spectral range of 350–1050 nm (quantum efficiency: 90% at 620 nm) was employed for photon detection and XY-imaging. To increase detection sensitivity, the variable binning was set to 2 × 2 resulting in final resolutions of 512 × 512 pixels and 26 × 26 µm² pixel size (slow scan mode). The exposure time was set to 60 s and the images were analyzed with the IndiGo software (Version 2.0.5.0, Berthold Technologies, Bad Wildbad, Germany). The presented images are a representative selection from the series of photos with the highest detected signal intensity level for each treatment. The experiment was repeated twice on different biological samples.

Hesari N., Szegő A., Mirmazloum I., Pónya Z., Kiss-Bába E., Kolozs H., Gyöngyik M., Vasas D, & Papp I. (2023). High-Nitrate-Supply-Induced Transcriptional Upregulation of Ascorbic Acid Biosynthetic and Recycling Pathways in Cucumber. Plants, 12(6), 1292.

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Transient Expression of nLUC/cLUC Fusions

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The LCI experiments were carried out as described previously (Chen et al., 2008 (link)). The nLUC/cLUC fusion plasmids and conjugative P19 plasmid were introduced into Agrobacterium strains GV3101 and EHA105, respectively. A single colony was transferred to Luria-Bertani medium and cultured overnight to OD₆₀₀ = 0.6–0.8. The culture was pelleted, washed twice with transformation buffer (10 mM MES [pH 5.6], and 10 mM MgCl₂), and resuspended to a final OD₆₀₀ of 1.5. Various nLUC/cLUC fusion constructs were mixed with an equal volume of P19. The bacteria were supplemented with 200 mM acetosyringone and incubated at 28°C for 3–5 h without shaking. The bacterial suspensions were infiltrated into fully expanded young N. benthamiana leaves with a needleless syringe. The plants were grown for 2 d under LD conditions. The infiltrated leaves were sprayed with 2 μM luciferase (dissolved in 0.02% Triton X-100) and incubated in the dark for 10 min before imaging. Luminescence was captured using a NightSHADE LB985 plant imaging system equipped with a CCD camera (Berthold Technologies). The experiments were repeated at least three times.

Zha P., Liu S., Li Y., Ma T., Yang L., Jing Y, & Lin R. (2019). The Evening Complex and the Chromatin-Remodeling Factor PICKLE Coordinately Control Seed Dormancy by Directly Repressing DOG1 in Arabidopsis. Plant Communications, 1(2), 100011.

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Protein-Protein Interaction Assays in Planta

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For the LCI assay, the CDSs of TaD14-4D and 4D-HapII were cloned into n-LUC vectors and the CDS of TaMAX2 was cloned into c-LUC vector to generate the TaD14-4D–n-LUC, 4D-HapII–nLUC, and TaMAX2–c-LUC constructs.
For the BiFC assay, the CDSs of the TaD14-4D and 4D-HapII were fused with N-terminal YFP, and TaMAX2 and TaD53 were fused with C-terminal YFP.
Different combinations of the above recombinant constructs were coinfiltrated into Nicotiana benthamiana leaves by Agrobacterium tumefaciens mediated transformation. The corresponding empty vectors were used as negative controls. The LUC and YFP signals were observed 48 72 h after infiltration by the Night SHADE LB 985 Plant Imaging System (Berthold Technologies, Bad Wildbad, Germany) and confocal microscope (LSM880; Carl Zeiss, Oberkochen, Germany).

Liu R., Hou J., Li H., Xu P., Zhang Z, & Zhang X. (2021). Association of TaD14-4D, a Gene Involved in Strigolactone Signaling, with Yield Contributing Traits in Wheat. International Journal of Molecular Sciences, 22(7), 3748.

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Protein-protein interaction analysis using yeast two-hybrid and LCI assays

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The yeast two‐hybrid assay was carried out as described previously (Wu and Li, 2017). The TaDA1‐A and TaGW2‐B CDSs were amplified and sub‐cloned into the pGBKT7 and pGADT7 vectors, respectively, to generate the TaDA1‐A‐BD and TaGW2‐B‐AD plasmids. After the co‐transformation of both plasmids into yeast AH109 cells, the interaction between the expressed proteins was determined by the growth of the co‐transformants on a selection medium (SD/‐Trp/‐Leu/‐His/‐Ade), following the Yeast Protocols Handbook (Takara Bio).
The LCI assay for the interaction between TaDA1‐A and TaGW2‐B was performed in N. benthamiana leaves as described previously (Liu et al., 2017). The full‐length TaDA1‐A and TaGW2‐B CDSs were fused with the N‐terminal and C‐terminal regions of the LUC reporter gene, respectively (TaDA1‐A‐nLUC and cLUC‐TaGW2‐B), and transformed into Agrobacterium tumefaciens strain GV3101. Agrobacteria harbouring the nLUC and cLUC derivative constructs were co‐infiltrated into N. benthamiana, and the LUC activity was imaged and analysed 48–72 h after infiltration using the NightSHADE LB 985 Plant Imaging System (Berthold Technologies, Bad Wildbad, Germany).

Liu H., Li H., Hao C., Wang K., Wang Y., Qin L., An D., Li T, & Zhang X. (2019). TaDA1, a conserved negative regulator of kernel size, has an additive effect with TaGW2 in common wheat (Triticum aestivum L.). Plant Biotechnology Journal, 18(5), 1330-1342.

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Bimolecular Fluorescence Complementation Assay

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GF14b was inserted into pCAMBIA‐split‐nLUC vector to generate GF14b‐nLUC fused vector. OsFTL12 and Hd3a were inserted into the pCAMBIA‐spit‐cLUC to generate cLUC‐OsFTL12 and cLUC‐Hd3a fused vector, respectively. The coding sequences of Hd3a and OsFTL12 were cloned into pCAMBIA1300 with the 35 S promoter as competitors. Constructed vectors were transformed into Agrobacterium strains EHA105. To detect whether Hd3a affects GF14b‐OsFTL12 interaction, EHA105 strains with recombinant GF14b‐nLUC, cLUC‐OsFTL12 and different concentrations of Hd3a competitors (100%, 200% and 300%) were mixed and co‐transformed into N. benthamiana leaves. To detect whether OsFTL12 affects the GF14b‐Hd3a interaction, EHA105 strains with GF14b‐nLUC, cLUC‐Hd3a and different concentrations of OsFTL12 competitors (100%, 200% and 300%) were mixed and co‐transformed into N. benthamiana leaves. Three leaves from independent N. benthamiana plants were infiltrated. After 48 h at 24 °C, 100 μm Luciferin was sprayed onto leaves, and LUC images were observed in the Nightshade LB985 plant imaging system (Berthold, Germany).

Zheng R., Meng X., Hu Q., Yang B., Cui G., Li Y., Zhang S., Zhang Y., Ma X., Song X., Liang S., Li Y., Li J., Yu H, & Luan W. (2023). OsFTL12, a member of FT‐like family, modulates the heading date and plant architecture by florigen repression complex in rice. Plant Biotechnology Journal, 21(7), 1343-1360.

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Transcriptional Activity Assay in N. benthamiana

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The transcriptional activity in Nicotiana benthamiana leaves was examined as described previously (Sun et al., 2012). The 2‐kb OsMIR530 promoter sequence was ligated to the luciferase reporter gene LUC in the plant binary vector pGWB35 (Nakagawa et al., 2007) via Gateway reactions (Invitrogen) to generate the reporter construct. To prepare the effector construct, the OsPIL15 coding sequence was cloned into the pCAMBIA 1300‐FLAG vector between the BamHI and SpeI restriction sites. The reporter and effector constructs were inserted separately into A. tumefaciens strain GV3101 cells for the subsequent co‐infiltration of N. Benthamiana leaves. The LUC signals were detected and quantified with the NightSHADE LB 985 Plant Imaging System (Berthold, Bad Wildbad, Germany) at 48 h after infiltrations. Ten independent N. benthamiana leaves were infiltrated and analyzed for each of three biological replicates. Details regarding the primers used for this assay are listed in Table S1.

Sun W., Xu X.H., Li Y., Xie L., He Y., Li W., Lu X., Sun H, & Xie X. (2020). OsmiR530 acts downstream of OsPIL15 to regulate grain yield in rice. The New Phytologist, 226(3), 823-837.

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Luciferase Tagging and Agroinfiltration

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The full-length CDS of SiUBC32, SiBRI1 or SiSGD1 were cloned into the pCAMBIA-1300-cLUC or pCAMBIA-1300-nLUC binary vectors, to generate N-terminus (nLUC) or C-terminus (cLUC) fused and truncated luciferase tags on the target proteins respectively. All constructs were paired and transformed into Agrobacterium strain GV3101 (pSoup-p19) for agroinfiltration of N. benthamiana leaves. After 36–48 h of co-infiltration, the luciferase substrate D-luciferin (Promega, Cat # P1043) was sprayed onto the leaf surface. The chemiluminescence signal was observed using the NightSHADE LB 985 plant imaging system (Berthold Technologies, Germany).

Tang S., Zhao Z., Liu X., Sui Y., Zhang D., Zhi H., Gao Y., Zhang H., Zhang L., Wang Y., Zhao M., Li D., Wang K., He Q., Zhang R., Zhang W., Jia G., Tang W., Ye X., Wu C, & Diao X. (2023). An E2-E3 pair contributes to seed size control in grain crops. Nature Communications, 14, 3091.

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Agrobacterium-mediated Transient Assays in N. benthamiana

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All constructs for LCI and BiFC assays were individually transformed into Agrobacterium tumefaciens strain EHA105 and then infiltrated into the leaves of N. benthamiana with different combinations as indicated in the figures. For BiFC assay, the N. benthamiana leaves were co‐infiltrated with a combination of Agrobacterium tumefaciens strain EHA105 carrying the indicated plasmid pairs and the 35S::mRFP‐AHL22 plasmid (Xiao et al., 2009). Samples were incubated in darkness for 24 h after the infiltration and then transferred to WL conditions (16‐h light/8‐h dark) for 24 to 36 h. For imaging the luciferase luminescence, the leaves were detached and photographed using the NightShade LB985 Plant Imaging System (Berthold Technologies) with a 60 s exposure time, 4 × 4 binning, slow readout and high gain after spraying with 20 mg/mL potassium luciferin (Gold Biotech, USA). For imaging the fluorescence of the reunion in BiFC, the N. benthamiana leaves were observed under confocal microscopy (Zeiss LSM710). All these experiments were independently repeated at least three times.

Li Q., Wu G., Zhao Y., Wang B., Zhao B., Kong D., Wei H., Chen C, & Wang H. (2020). CRISPR/Cas9‐mediated knockout and overexpression studies reveal a role of maize phytochrome C in regulating flowering time and plant height. Plant Biotechnology Journal, 18(12), 2520-2532.

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