Journal: Planta

Article Title: Absence of photosynthetic state transitions in alien chloroplasts

doi: 10.1007/s00425-019-03187-2

Figure Lengend Snippet: Isolation of light-harvesting complexes. a Thylakoid protein preparations separated on the basis of pI by isoelectric focusing. Photographs of representative gels from tobacco (T), the Nt ( Hn ) cybrid and henbane (H). The boxed area near the cathode defines the site where thylakoids were loaded into the gel. The pH scale and distance from cathode are marked above and below the gel, respectively. The former is approximate as pH gradients varied slightly between gels. Images have been cropped to show the central aspect of the gel tray for easy comparison. b SDS–PAGE separation of LHCII polypeptides from the broad IEF band in a 15% acrylamide gel. The protein marker is shown in the first lane (m) the standard weights are indicated. The pink arrow indicates the novel band in the Nt ( Hn ) separation. c Density plots of the MW profiles shown in b . The plots from left to right correspond to the top to bottom of the gel in b , respectively. Tobacco is shown in green and the Nt ( Hn ) cybrid in blue. Peaks were numbered 1–7 in tobacco with equivalent peaks in Nt ( Hn ) density plots assigned letters ( A – H )

Article Snippet: Two IEF electrode strips (GE Healthcare) were saturated, one in anode solution (5.3% (v/v) H 3 PO 4 ) and one in cathode solution (1 M NaOH), and carefully positioned at the anode and the cathode positions, respectively.

Techniques: Isolation, SDS Page, Acrylamide Gel Assay, Marker

Journal: Frontiers in Immunology

Article Title: PD-1 knockout on cytotoxic primary murine CD8 + T cells improves their motility in retrovirus infected mice

doi: 10.3389/fimmu.2024.1338218

Figure Lengend Snippet: Genetic editing of primary CTLs for the evaluation of CTL motility and function. (A) Schematic overview of the workflow. Gene edited CTLs were generated from naïve CD8+ T cells obtained from the blood of TCR-Lck-tdTom mice using the Lonza 4D nucleofector system. C57BL/6 and DEREG recipient mice were infected with FV and subsequently received 1,000 CTLs, which were treated with gRNA/Cas9 RNPs targeting PDCD1 or unspecific gRNA/Cas9 RNPs (control) or unnucleofected primary naïve CTLs. Intravital two-photon bone marrow microscopy and flow cytometry were performed at 14 dpi. Schematic overview was created with BioRender.com. (B) Frequencies of PD-1 expressing PDCD1 -targeted, control and unnucleofected CD8+ T cells of the in-vitro KO validation (median ± IQR). Data was obtained for each single nucleofected sample in 3 independent experiments. (C) Frequencies of PD-1 expressing transferred or endogenous activated CTLs of FV infected mice transferred with PDCD1 -targeted, control and unnucleofected CTLs at 14 dpi (median ± IQR). Data were obtained in 1-4 independent experiments with 1-4 mice. P-values were obtained by Kruskal-Wallis test followed by a corrected Dunn’s multiple comparison test. *p ≤ 0.05.

Article Snippet: After incubation 1 µL electroporation enhancer (IDT, cat. No. 1075916) was added to the solution and isolated CD8+ T cells were resuspended in 20 µL freshly prepared P3 buffer, added to the 5 µL gRNA/Cas9-RNP mix and transferred to the bottom hole of a well of the Lonza nucleofector strip.

Techniques: Generated, Infection, Microscopy, Flow Cytometry, Expressing, In Vitro, Comparison