Journal: bioRxiv

Article Title: Production and secretion of functional full-length SARS-CoV-2 spike protein in Chlamydomonas reinhardtii

doi: 10.1101/2021.12.13.472433

Figure Lengend Snippet: Design of constructs for the production and secretion of the SARS CoV-2 spike protein in Chlamydomonas. (a) Schematic representation of the two level 0 parts encoding the spike protein. The codon adapted coding region is depicted by black boxes and incorporated RBSC2 introns by thin lines. The braces highlight regions modified by site directed mutagenesis and the resulting amino acid modifications. Colored boxes represent fusion sites according to the MoClo syntax. (b) Level 2 MoClo devices consisting of a level 1 module containing the aadA resistance marker under control of the PSAD promoter and terminator (S R ), and level 1 transcriptional units for the expression of the spike protein. The latter contain the HSP70A-RBCS2 promoter ( P AR ), sequences encoding various secretion signals (cCA, ARS, GLE, sec), the two parts of the spike protein (CoV2-S up and down), a glycomodule of 20 serine/proline repeats (SP20) fused to the HA epitope and/or an octa-his tag, and the RPL23 terminator ( T RPL23 ). (c) Production and secretion of the spike protein in transformants generated with the five level 2 devices listed in (b). Shown are representative immunoblots detecting HA-tagged proteins in cell lysates (CL) (corresponding to 2 μg chlorophyll) and 1.7 mL of culture medium (CM) after TCA precipitation. The asterisk indicates a cross reaction of the HA antibody. (d) Three independent preparations of spike protein secreted by a S-GSAS/PP transformant into 1.7 mL culture medium (CM) and precipitated with TCA, and increasing amounts of purified recombinant mCherry carrying a C-terminal HA tag were analyzed by immunoblotting using an HA antibody.

Article Snippet: Synthesis and cloning into the pUC57 vector were done by BioCat (Heidelberg), yielding level 0 part pMBS514 ( SP20-3xHA ).

Techniques: Construct, Modification, Mutagenesis, Marker, Expressing, Generated, Western Blot, TCA Precipitation, Purification, Recombinant

Journal: International Journal of Physiology, Pathophysiology and Pharmacology

Article Title: ZO-1 associates with α3 integrin and connexin43 in trabecular meshwork and Schlemm’s canal cells

doi:

Figure Lengend Snippet: RT-PCR showing detection of α3 and β1 integrin mRNA transcripts in trabecular meshwork cells.

Article Snippet: Monoclonal antibodies against α3 integrin (Cat. No. NBP1-97692), and β1 integrin (NBP2-22191) were obtained from Novus Biologicals Laboratories (Littleton, CO, USA).

Techniques: Reverse Transcription Polymerase Chain Reaction

Journal: International Journal of Physiology, Pathophysiology and Pharmacology

Article Title: ZO-1 associates with α3 integrin and connexin43 in trabecular meshwork and Schlemm’s canal cells

doi:

Figure Lengend Snippet: Western Blotting showing detection of α3 integrin and β1 integrin in trabecular meshwork cells. The weak lower molecular weight bands in α3 integrin immunoblotting may correspond the degradation products of α3 integrin protein.

Article Snippet: Monoclonal antibodies against α3 integrin (Cat. No. NBP1-97692), and β1 integrin (NBP2-22191) were obtained from Novus Biologicals Laboratories (Littleton, CO, USA).

Techniques: Western Blot, Molecular Weight

Journal: International Journal of Physiology, Pathophysiology and Pharmacology

Article Title: ZO-1 associates with α3 integrin and connexin43 in trabecular meshwork and Schlemm’s canal cells

doi:

Figure Lengend Snippet: Partial co-localization of α3 integrin with ZO-1 in TM cells. Double immunofluorescence labeling showing α3 integrin (red, A) and ZO-1 (green, C) with partial co-localization in overlay (yellow, E) in TM cells, while β1 integrin (red, B) and ZO-1 (green, D) showing lack of co-localization in overlay (F).

Article Snippet: Monoclonal antibodies against α3 integrin (Cat. No. NBP1-97692), and β1 integrin (NBP2-22191) were obtained from Novus Biologicals Laboratories (Littleton, CO, USA).

Techniques: Immunofluorescence, Labeling

Journal: International Journal of Physiology, Pathophysiology and Pharmacology

Article Title: ZO-1 associates with α3 integrin and connexin43 in trabecular meshwork and Schlemm’s canal cells

doi:

Figure Lengend Snippet: Partial co-localization of α3 integrin with ZO-1 in SC cells. Double immunofluorescence labeling showing α3 integrin (red, A) and ZO-1 (green, C) with partial co-localization in overlay (yellow, E) in SC cells. Immunofluorescence labeling of CD31 (red) in SC cells (B), but there is no CD31 labeling in TM cells (D), while Cx43 (red) and ZO-1 (green) showing co-localization in overlay in TM cells (F).

Article Snippet: Monoclonal antibodies against α3 integrin (Cat. No. NBP1-97692), and β1 integrin (NBP2-22191) were obtained from Novus Biologicals Laboratories (Littleton, CO, USA).

Techniques: Immunofluorescence, Labeling

Journal: International Journal of Physiology, Pathophysiology and Pharmacology

Article Title: ZO-1 associates with α3 integrin and connexin43 in trabecular meshwork and Schlemm’s canal cells

doi:

Figure Lengend Snippet: Co-IP of ZO-1 with α3 integrin and Cx43 in TM lysates. Immunoblots of IP material using anti-α3 integrin probed with anti-ZO-1 antibody revealed the presence of ZO-1 (lane 2), which co-migrated with ZO-1 in lysates of PTM cells (lane 1) and from IP materials using anti-Cx43 as positive controls for ZO-1 detection (lane 3). ZO-1 was absent from IP material after omission of anti-α3 integrin during the IP procedure, which was included as a negative control (lane 4).

Article Snippet: Monoclonal antibodies against α3 integrin (Cat. No. NBP1-97692), and β1 integrin (NBP2-22191) were obtained from Novus Biologicals Laboratories (Littleton, CO, USA).

Techniques: Co-Immunoprecipitation Assay, Western Blot, Negative Control

Journal: Molecular Cell

Article Title: The TRESLIN-MTBP complex couples completion of DNA replication with S/G2 transition

doi: 10.1016/j.molcel.2022.08.006

Figure Lengend Snippet:

Article Snippet: GAPDH , Novus Biologicals , Cat# NB600-502.

Techniques: Recombinant, Western Blot, Microscopy, Mutagenesis, Software, Cell Culture, Transfection, Protease Inhibitor

Journal: Nature communications

Article Title: MEIOB exhibits single-stranded DNA-binding and exonuclease activities and is essential for meiotic recombination

doi: 10.1038/ncomms3788

Figure Lengend Snippet: ( a ) Identification of MEIOB-associated proteins from P18 testes by immunoprecipitation and mass spectrometry. The gel was stained with SYPRO Ruby. ( b ) Co-immunoprecipitation of MEIOB with RPA2 and SPATA22 from testicular protein extracts. The asterisk indicates a non-specific band. ( c ) Inter-dependent localization of MEIOB and SPATA22 on meiotic chromosomes from spermatocytes. Spata22 −/− refers to Spata22 repro42/repro42 mice . Scale bar, 10 µm. ( d ) Western blot analysis of MEIOB and SPATA22 proteins in testes from wild type, Meiob −/− , and Spata22 −/− mutant mice.

Article Snippet: The following primary antibodies were used for immunofluorescence on spread nuclei: SYCP1 (1:50, Cat. No. ab15090, Abcam), SYCP2 , SYCP3 (1:100, Cat. No. 611230, BD Biosciences), γH2AX (1:500, Cat. No. 16-202A, Clone JBW301, Millipore), ATR (1:200, Cat. No. PC538, EMD Biosciences), RPA1 (1:50, Cat. No. ab87272, Abcam), RPA2 (1:100, Cat. No. 2208S, clone 4E4, Cell Signaling Technology), RAD51 (1:50, gift from the late Peter Moens), DMC1 (1:50, Cat. No. sc-22768 (H-100), Santa Cruz Biotech), MLH1 (1:50, Cat. No. 550838, clone G168-15, BD Biosciences), MSH4 (1:50, gift from Chengtao Her) , SPATA22 (1:50, Cat. No. 16989-1-AP, ProteinTech Group), and TEX11 (1:10) .

Techniques: Immunoprecipitation, Mass Spectrometry, Staining, Western Blot, Mutagenesis

Journal: Nature communications

Article Title: MEIOB exhibits single-stranded DNA-binding and exonuclease activities and is essential for meiotic recombination

doi: 10.1038/ncomms3788

Figure Lengend Snippet: Key steps of meiotic recombination are illustrated. We propose that the RPA-MEIOB-SPATA22 complex coats both the D-loop and the ssDNA of the second end. In this model, the interaction between RPA and MEIOB-SPATA22 mediates second end capture. Most DSBs are repaired through the SDSA (synthesis dependent strand annealing) pathway, in which the D-loop collapses back to its sister chromatid. After second end capture, intermediates continue to form double Holliday junctions, which are resolved into either crossovers or non-crossovers. We propose that MEIOB removes the 3’ flaps resulting from the first end DNA synthesis in both dHJ and SDSA pathways.

Article Snippet: The following primary antibodies were used for immunofluorescence on spread nuclei: SYCP1 (1:50, Cat. No. ab15090, Abcam), SYCP2 , SYCP3 (1:100, Cat. No. 611230, BD Biosciences), γH2AX (1:500, Cat. No. 16-202A, Clone JBW301, Millipore), ATR (1:200, Cat. No. PC538, EMD Biosciences), RPA1 (1:50, Cat. No. ab87272, Abcam), RPA2 (1:100, Cat. No. 2208S, clone 4E4, Cell Signaling Technology), RAD51 (1:50, gift from the late Peter Moens), DMC1 (1:50, Cat. No. sc-22768 (H-100), Santa Cruz Biotech), MLH1 (1:50, Cat. No. 550838, clone G168-15, BD Biosciences), MSH4 (1:50, gift from Chengtao Her) , SPATA22 (1:50, Cat. No. 16989-1-AP, ProteinTech Group), and TEX11 (1:10) .

Techniques: DNA Synthesis