GFP-Tag Rabbit Polyclonal Antibody - #T0006

FIGURE 1| Interaction of OsHRZ1 with OsPRI2 and OsPRI3..(d) Degradation of OsPRI2 or OsPRI3 was carried out by detecting the OsPRI2/3‐GFP protein level in co‐infiltration experiments with increasing amounts of OsHRZ1‐GFP. GFP proteins were used as an internal control. Anti‐GFP antibody was used in western blot. Protein molecular weight (in kDa) is indicated. Stars indicate the non‐specific bands. Numbers indicate the ratio of the concentrations of agrobacteria used in co‐infiltration. Empty vector, a binary vector pOCA30 with a 35S promoter; GFP, 35S:GFP in pOCA30; OsPRI2‐GFP, 35S:OsPRI2‐GFP in pOCA30; OsPRI3‐GFP, 35S:OsPRI3‐GFP in pOCA30; OsHRZ1‐GFP, 35S:OsHRZ1‐GFP in pOCA30

FIGURE 1 Interaction of OsHRZ1 with OsPRI2 and OsPRI3. (a) Yeast two‐hybrid assays. Yeast co‐transformed with different BD and AD plasmid combinations were spotted in parallel in 10‐fold dilution series on synthetic dropout medium lacking Leu/Trp/His/Ade. The C‐terminal truncated OsHRZ1 and full‐length OsPRI2/3 were cloned into pGBKT7 and pGADT7, respectively. OsHRZ1‐C/OsPRI1, positive control. OsHRZ1‐ C/Empty, negative control. (b) Pull‐down assay. OsHRZ1 was fused with the GST tag and OsPRI2/3 were fused with the His tag. Recombinant proteins were expressed in E. coli. Proteins were pulled down by glutathione Sepharose 4B and detected using the anti‐His antibody. Protein molecular weight (in kDa) is indicated. (c) CoIP assay. Total proteins from different combinations with OsHRZ1‐GFP and HA‐OsPRI2/HA‐OsPRI3/ HA‐GUS were immunoprecipitated with GFP‐Trap followed by immunoblotting with the indicated antibodies. HRZ1‐GFP/HA‐GUS, negative control. Protein molecular weight (in kDa) is indicated. (d) Degradation of OsPRI2 or OsPRI3 was carried out by detecting the OsPRI2/3‐GFP protein level in co‐infiltration experiments with increasing amounts of OsHRZ1‐GFP. GFP proteins were used as an internal control. Anti‐GFP antibody was used in western blot. Protein molecular weight (in kDa) is indicated. Stars indicate the non‐specific bands. Numbers indicate the ratio of the concentrations of agrobacteria used in co‐infiltration. Empty vector, a binary vector pOCA30 with a 35S promoter; GFP, 35S:GFP in pOCA30; OsPRI2‐GFP, 35S:OsPRI2‐GFP in pOCA30; OsPRI3‐GFP, 35S:OsPRI3‐GFP in pOCA30; OsHRZ1‐GFP, 35S:OsHRZ1‐GFP in pOCA30. (e) Cell‐free degradation. Ten‐day‐old roots grown in Fe‐sufficient solution were harvested and used for protein extraction. Incubation with or without MG132 was performed over the indicated time course

Figure 1. Nucleocytoplasmic trafficking of M protein promotes the replication and cytopathogenicity of NDV by affecting viral RNA synthesis and transcription. (A) The subcellular localization of M protein in rSS1GFP- and rSS1GFP-M/NLSm-infected BSR-T7/5 cells at 6, 12, 18 and 24 hpi. DAPI was used to stain nuclei. Original magnification was 1 × 200. (B) Virus titers were detected in BSR-T7/5 cells at the indicated time points. (C) The CPE and GFP were observed in virus-infected BSR-T7/5 cells at 12 and 24 hpi. Original magnification was 1 × 200. (D) The viral RNA synthesis corresponding to the NP and P genes and (E) viral transcription corresponding to the M and GFP genes in rSS1GFP- and rSS1GFP-M/NLSm-infected BSR-T7/5 cells were detected by qRT-PCR. (F) The expression levels of NP, M and GFP proteins in rSS1GFP- and rSS1GFP-M/NLSm-infected BSR-T7/5 cells were examined by Western blotting. The relative levels of the NP, M and GFP proteins were compared with the control GAPDH expression. Each data indicates the mean ± SD of three independent experiments. P-values are indicated by asterisks (*P < 0.05, **P < 0.01, ***P < 0.001 compared to the value of rSS1GFP-M/NLSm).

Figure 2. |Transgene expression after intracavernosal AAV injection. Panel A shows the representative immunofluorescence images(magnification  100) of GFP protein at 4 weeks after intracavernosal delivery of AAV-GFP