Journal: Molecular and Cellular Biochemistry

Article Title: Large DNA fragment knock-in and sequential gene editing in Plasmodium falciparum : a preliminary study using suicide-rescue-based CRISPR/Cas9 system

doi: 10.1007/s11010-023-04711-5

Figure Lengend Snippet: The expression analysis of the integrated K119OD/R787OD recombinant protein. A RT-PCR detection of the transcript of the integrated k119od gene (left) and Western blot analysis of its coding protein (right). V5, V5 epitope; GAPDH (glyceraldehyde 3-phosphate dehydrogenase), internal reference. B RT-PCR detection of the transcript of the integrated r787od gene (left) and Western blot analysis of its coding protein (right). C Indirect immunofluorescence microscopy of K119OD (left) and R787 (right) transgenic parasites. 3D7WT, wild-type Pf 3D7 parasite, negative control. Scale bar, 10 μm

Article Snippet: Parasite samples were collected by Percoll gradient [ ] and Western blot was performed routinely; mouse anti-V5 antibody [Code No: D199986; Sangon Biotech (Shanghai) Co., Ltd.] and horseradish peroxidase-conjugated goat anti-mouse secondary antibody (Code No: 31430; Invitrogen) were used.

Techniques: Expressing, Recombinant, Reverse Transcription Polymerase Chain Reaction, Western Blot, Immunofluorescence, Microscopy, Transgenic Assay, Negative Control

Journal: Molecular and Cellular Biochemistry

Article Title: Large DNA fragment knock-in and sequential gene editing in Plasmodium falciparum : a preliminary study using suicide-rescue-based CRISPR/Cas9 system

doi: 10.1007/s11010-023-04711-5

Figure Lengend Snippet: The validation of obtained episome-free parasites. A PCR detection of episomal pCBS-yFCU- pfs47 residual. bsd gene carried by the plasmid was detected (396 bp) and kahrp gene was employed as an internal reference (2353 bp). d5, 3D7R787OD parasites under negative selection using 5-FC initiated at day 5 after blasticidin S withdrawn and last up to 4 weeks; d15, 3D7R787OD parasites under negative selection using 5-FC initiated at day 15 after blasticidin S was withdrawn and last less than 3 weeks; + BS, 3D7R787OD parasites maintained continuously with blasticidin S (positive control); – BS, 3D7R787OD parasites maintained without blasticidin S; ddw, double distilled water (blank control); wt, wild-type Pf 3D7 parasites (negative control). B Indirect immunofluorescence microscopy of episome-free R787 transgenic parasites (3D7R787ODep − ) labeled using V5-tag antibody and Cy3-conjugated secondary antibody, wild-type Pf 3D7 parasites were used as negative control. BF, bright field. Scale bar, 10 μm

Article Snippet: Parasite samples were collected by Percoll gradient [ ] and Western blot was performed routinely; mouse anti-V5 antibody [Code No: D199986; Sangon Biotech (Shanghai) Co., Ltd.] and horseradish peroxidase-conjugated goat anti-mouse secondary antibody (Code No: 31430; Invitrogen) were used.

Techniques: Biomarker Discovery, Plasmid Preparation, Selection, Positive Control, Control, Negative Control, Immunofluorescence, Microscopy, Transgenic Assay, Labeling

Journal: bioRxiv

Article Title: Nucleic acid sensing by STING induces an interferon-like antiviral response in a marine invertebrate

doi: 10.1101/2022.12.18.520954

Figure Lengend Snippet: (A) Schematic representations of various deletion mutants of LvSTING, LvIRF and LvIKKε. (B) Interactions between GFP-tagged LvIRF and HA-tagged full-length or truncated LvSTING were analyzed by Co-IP assays. (C) Interactions between HA-tagged LvSTING and GFP-tagged full-length or truncated LvIRF were analyzed by Co-IP assays. (D) Interactions between V5-tagged LvIKKε and HA-tagged full-length or truncated LvSTING were analyzed by Co-IP assays. (E) Interactions between HA-tagged LvSTING and V5-tagged full-length and truncated LvIKKε was analyzed by Co-IP assays. (F) Relative induction of the promoter activities of LvVago4 mediated by LvIRF-V5, LvIKKε-V5 and LvSTING-HA or LvSTING-1– 300-HA in Drosophila S2 cells treated by 2′3′-cGAMP using dual luciferase assays. Ectopically expressed proteins were detected by Western blotting. Actin was used as a protein loading control. The bars indicated the mean ± SD of the luciferase activities ( n = 3). The data was analyzed statistically by student’s t -test ( ** p < 0.01). All experiments were replicated three times with similar results.

Article Snippet: The supernatants (500 µL) were incubated with 30 µL of anti-GFP (MBL), anti-V5 (Sigma) or anti-HA affinity gel (Sigma) at 4 °C for 4 h. The agarose affinity gels were washed with PBS five times and subjected to SDS-PAGE assay and Western blotting using rabbit anti-GFP antibody (Sigma), rabbit anti-V5 antibody (Merck) or rabbit anti-HA antibody (Sigma).

Techniques: Co-Immunoprecipitation Assay, Luciferase, Western Blot