Log In Sign up
The largest database of trusted experimental protocols

Eif4g

Manufactured by Santa Cruz Biotechnology
Visit Supplier
Sourced in United States

EIF4G, also known as Eukaryotic translation initiation factor 4 gamma, is a protein that plays a crucial role in the initiation of protein synthesis. It is a component of the eukaryotic translation initiation complex, which is responsible for bringing the mRNA and the ribosome together to begin the translation process. EIF4G acts as a scaffold, binding to other translation initiation factors and facilitating their interactions, thus enabling the efficient recruitment and assembly of the translation machinery.

Automatically generated - may contain errors

Lab products found in correlation

Eif4e, by Santa Cruz Biotechnology (4 mentions) Gapdh, by Santa Cruz Biotechnology (4 mentions) Supersignal west pico plus chemiluminescent substrate, by Thermo Fisher Scientific (3 mentions) Normal rabbit igg, by Merck Group (2 mentions) β tubulin, by Cell Signaling Technology (2 mentions) Iblot 2 gel transfer device, by Thermo Fisher Scientific (2 mentions) Bolt reducing agent, by Thermo Fisher Scientific (2 mentions) Bolt lds buffer, by Thermo Fisher Scientific (2 mentions) Nucleolin, by Santa Cruz Biotechnology (2 mentions) G3bp1, by Santa Cruz Biotechnology (2 mentions) Tia 1, by Santa Cruz Biotechnology (2 mentions) Rack1, by Santa Cruz Biotechnology (2 mentions) β actin, by Proteintech (2 mentions) Phosphatase inhibitor cocktails 2 and 3, by Merck Group (2 mentions) Ab21060, by Abcam (1 mentions) Trans blot turbo transfer system, by Bio-Rad (1 mentions) Anti rabbit igg antibody, by Jackson ImmunoResearch (1 mentions) Mcl 1, by Santa Cruz Biotechnology (1 mentions) N myc, by Cell Signaling Technology (1 mentions) Protease inhibitor, by Merck Group (1 mentions)

13 protocols using eif4g

1

Antibody Production and Characterization

Check if the same lab product or an alternative is used in the 5 most similar protocols
The rabbit polyclonal anti-M2-1, anti-N and anti-M were obtained by repeated injection of purified recombinant protein produced in Escherichia coli as previously described42 (link). The mouse anti-cellular proteins antibodies used in western blotting were from Santa Cruz: PABP (10E10; 0.4 µg/ml), eIF4G (A-10; 0.4 µg/ml), eIF4E (P-2; 0.4 µg/ml), eIF3η (C-5; 0.4 µg/ml) and eIF4AI/II (H-5; 0.4 µg/ml). The Santa Cruz mouse anti-PABP antibody and an abcam rabbit anti-IMPDH2 antibody (ab75790) were used in co-immunoprecipitation at a concentration of 10 µg/ml. The rabbit anti-PABP (ab21060; 2 µg/ml) and mouse anti-M2-1 (ab94805; 2 µg/ml) used in immunofluorescence staining and Duolink were from Abcam. Secondary antibodies (2 µg/ml) raised against mouse or rabbit IgG (H + L) and conjugated to Alexa Fluor 488, 594 or 647 were from Invitrogen. Secondary antibodies (0.1 µg/ml) raised against mouse or rabbit IgG (H + L) and conjugated to horseradish peroxidase were from Promega.
Bouillier C., Cosentino G., Léger T., Rincheval V., Richard C.A., Desquesnes A., Sitterlin D., Blouquit-Laye S., Eléouët J.F., Gault E, & Rameix-Welti M.A. (2019). The Interactome analysis of the Respiratory Syncytial Virus protein M2-1 suggests a new role in viral mRNA metabolism post-transcription. Scientific Reports, 9, 15258.
+ Open protocol
+ Expand
2

Western Blot Analysis of Protein Expression

Check if the same lab product or an alternative is used in the 5 most similar protocols
Samples were collected using 1 × SDS buffer and run on 10% polyacrylamide gels. Gels were transferred in the Trans-Blot Turbo transfer system (Bio-Rad). Membranes were blocked using 5% BSA in 1× tris-buffered saline with 0.2% Tween (TBST) and probed with primary eIF4G (1:1,000; Santa Cruz Biotech), ODC1 (1:1,000; Invitrogen), SAT1 (1:1,000; Novus Biologicals), and b-actin (1:1,000; ProteinTech) antibody. Membranes were washed in 1× TBST and placed in secondary anti-mouse IgG (1:15,000; Jackson Immunoresearch) or anti-rabbit IgG antibody (1:15,000; Jackson Immunoresearch) and incubated at room temperature for 1h. Again, membranes were washed in 1× TBST, and SuperSignal West Pico Plus chemiluminescent substrate (Thermo-Fisher) was applied to membranes and developed on a molecular imager, Bio-Rad GelDoc XR1 imaging system (Bio-Rad). Images were quantified with ImageJ.
Mastrodomenico V., LoMascolo N.J., Firpo M.R., Villanueva Guzman M.D., Zaporowski A, & Mounce B.C. (2023). Persistent Coxsackievirus B3 Infection in Pancreatic Ductal Cells In Vitro Downregulates Cellular Polyamine Metabolism. mSphere, 8(3), e00036-23.
+ Open protocol
+ Expand
3

Immunoblot analysis of cellular signaling

Check if the same lab product or an alternative is used in the 5 most similar protocols
Cells were lysed in lysis buffer (25 mM Tris pH 8.0, 125 mM NaCl, 1 mM MgCl2, 1% Triton X-100, 1 mM sodium orthovanadate, 1 mM sodium fluoride, 1X protease inhibitor (Sigma-Aldrich), phosphatase inhibitor cocktails #2 and #3 (Sigma-Aldrich), and 1 mM PMSF). Cell lysates were resolved on a Nupage 4-12% Bis-Tris gradient gel (Life Technologies) and probed using antibodies recognising phospho- and total eIF4E1 (Cell Signaling #9741 and Santa Cruz sc-9976), phospho- and total ERK (#9101 and #9102, Cell Signaling), c-Myc (#9402), n-Myc (#9405), p-MNK1/2 (#2111), dicer (#3363), eIF4A (#2013) (Cell Signaling), GAPDH (ab8245), BTK (ab54219), YY1 (ab12132) (Abcam), eIF4E3 (Proteintech: N-terminal, #17282-1-AP), MNK1 (sc-6965), MNK2 (sc-6964), CDK6 (sc-7961), eIF4G (sc-11373), MCL-1 (#sc-819) (Santa Cruz) or GFP (EVN-AB513, Axxora). All primary antibodies were used at 1: 1,000 dilution. Densitometry analyses were performed using ImageJ software (NIH) and presented as ratio of target band signal intensity to GAPDH band signal intensity. Full immunoblots are presented in Supplementary Fig.10.
Landon A.L., Muniandy P.A., Shetty A.C., Lehrmann E., Volpon L., Houng S., Zhang Y., Dai B., Peroutka R., Mazan-Mamczarz K., Steinhardt J., Mahurkar A., Becker K.G., Borden K.L, & Gartenhaus R.B. (2014). MNKs act as a regulatory switch for eIF4E1 and eIF4E3 driven mRNA translation in DLBCL. Nature Communications, 5, 5413.
+ Open protocol
+ Expand
4

Antibody Validation for Immunoblotting and Immunoprecipitation

Cited 1 time
Check if the same lab product or an alternative is used in the 5 most similar protocols
The following primary antibodies utilized throughout this study for immunoblotting or immunoprecipitation: CAP-D3 (Bethyl Laboratories), CAP-H2 (Bethyl Laboratories), EPRS (Bethyl Laboratories), L13a (Cell Signaling Technology), NSAP1 (AnaSpec Inc), GAPDH (Santa Cruz), eIF4G (Santa Cruz), eIF3e (Abcam), eIF4E (Abcam), IFN-γ (Biosource), Actin (Millipore), RNA Polymerase II (Abcam), β-tubulin (Cell Signaling Technology), Normal Rabbit IgG (Millipore). Purified polyclonal α-ORF1p was generated by OpenBiosystems and characterized by the laboratory of John Moran (University of Michigan School of Medicine[57 (link)]). EPRS WHEP-linker antibody was previously described [118 (link)].
Ward J.R., Vasu K., Deutschman E., Halawani D., Larson P.A., Zhang D., Willard B., Fox P.L., Moran J.V, & Longworth M.S. (2017). Condensin II and GAIT complexes cooperate to restrict LINE-1 retrotransposition in epithelial cells. PLoS Genetics, 13(10), e1007051.
+ Open protocol
+ Expand
5

Western Blot Analysis of Protein Expression

Check if the same lab product or an alternative is used in the 5 most similar protocols
Samples were collected with Bolt LDS Buffer and Bolt Reducing Agent (Invitrogen, Waltham, MA, USA) and run on polyacrylamide gels. Gels were transferred using the iBlot 2 Gel Transfer Device (Invitrogen). Membranes were probed with primary antibodies for eIF4G, (1:1000, Santa Cruz Biotechnology, Dallas, TX, USA), GAPDH (1:1000, Santa Cruz Biotechnology), and β-actin (1:5000, Santa Cruz Biotechnology). Membranes were treated with SuperSignal West Pico PLUS Chemiluminescent Substrate (ThermoFisher Scientific) and visualized on FluorChem E imager (Protein Simple, San Jose, CA, USA).
Dial C.N., Tate P.M., Kicmal T.M, & Mounce B.C. (2019). Coxsackievirus B3 Responds to Polyamine Depletion via Enhancement of 2A and 3C Protease Activity. Viruses, 11(5), 403.
+ Open protocol
+ Expand
6

Protein Distribution in Stress Granules

Check if the same lab product or an alternative is used in the 5 most similar protocols
Full table and dilutions for each antibody used can be found in Supplementary Table 4. YB-1 (Abcam: Ab12148), G3BP1 (Santa Cruz: sc81940), TIA1 (Santa Cruz: sc-1751), TIAR (Santa Cruz: sc-1749), eIF4G (Santa Cruz: sc-11373), Nucleolin (Santa Cruz: sc-9893), eIF4E (Santa Cruz: sc9976), Fus/TLS (Protein Tech Group: 11570-1-AP), TDP43 (Protein Tech Group: 10782-2-AP), Vigilin (Santa Cruz: 2404C3a), DHX36 (Protein Tech Group: 13159-1-AP), and GRSF1 (Aviva: ARP40382-P050).
Lyons S.M., Gudanis D., Coyne S.M., Gdaniec Z, & Ivanov P. (2017). Identification of functional tetramolecular RNA G-quadruplexes derived from transfer RNAs. Nature Communications, 8, 1127.
+ Open protocol
+ Expand
7

Western Blot Analysis of Translation Factors

Check if the same lab product or an alternative is used in the 5 most similar protocols
Proteins from cell lysates were separated by SDS-PAGE and probed with the indicated antibodies. Commercial antibodies were used to detect eIF4E (Transduction laboratories), RACK1, eIF2α, eIF4G (Santa Cruz), eEF2 (Cell Signaling), eIF4B, and Gemin5 (Novus). The ribosomal proteins P0, P1 and P2 were detected with the monoclonal antibody 3BH548 (link). Appropriate secondary antibodies (Thermo-Fisher) were used according to the manufacturer instructions. Protein signals were visualized with ECL plus (Millipore). Quantification of the signal detected was done in the linear range of the antibodies.
Lozano G., Francisco-Velilla R, & Martinez-Salas E. (2018). Ribosome-dependent conformational flexibility changes and RNA dynamics of IRES domains revealed by differential SHAPE. Scientific Reports, 8, 5545.
+ Open protocol
+ Expand
8

Analysis of Viral Protein Interactions Using Affinity Capture

Check if the same lab product or an alternative is used in the 5 most similar protocols
Actinomycin D, dextran sulfate and streptavidin were from Sigma. 5EU was from Invitrogen. Formaldehyde was from Polysciences. AZ27 was obtained from AstraZeneca R&D Boston30 (link), 31 (link). Stock solutions of Actinomycin D (1 mg/ml), streptavidin (1 mg/ml), dextran sulfate (40%, w/v) and 5EU (100 mM) were prepared in water. Formaldehyde (4%, v/v) was prepared in phosphate-buffered saline (PBS). AZ27 1 mM was stored in dimethyl sulfoxide and working dilution was prepared at 100 μM in MEM FCS 2% extemporaneously. The rabbit polyclonal anti-N was obtained by repeated injection of purified recombinant protein produced in Escherichia coli as described in ref. 55 (link). The mouse anti-cellular proteins antibodies: PABP (10E10; 1 µg/ml), eIF4G (A-10; 1 µg/ml), S6 (C-6; 1 µg/ml), L4 (RQ-7; 2 µg/ml) were from Santa Cruz. The rabbit anti-eRF1 (GTX 108271; 2 µg/ml) was from Gene Tex, the goat anti-TIA-1 (C-20; 2 µg/ml) was from Santa Cruz and the mouse anti-G3BP (2F3; 5 µg/ml) was from Sigma. Secondary antibodies (2 µg/ml) raised against mouse, rabbit or goat IgG (H + L) and conjugated to Alexa Fluor 488, 594 or 647 were from Invitrogen.
Rincheval V., Lelek M., Gault E., Bouillier C., Sitterlin D., Blouquit-Laye S., Galloux M., Zimmer C., Eleouet J.F, & Rameix-Welti M.A. (2017). Functional organization of cytoplasmic inclusion bodies in cells infected by respiratory syncytial virus. Nature Communications, 8, 563.
+ Open protocol
+ Expand
9

Western Blot and Immunofluorescence Antibodies

Check if the same lab product or an alternative is used in the 5 most similar protocols
The following antibodies were used for Western blotting and immunofluorescence: YB-1 (Abcam: Ab12148, Proteintech Group: 20339-1-AP, Bethyl Laboratories, Inc (A): A404-230-T, Bethyl Laboratories, Inc (B): A303-231-T), Dach1 (Proteintech Group: 10914-1-AP), Caprin1 (Proteintech Group: 14112-1-AP), G3BP1 (Santa Cruz Biotechnology, Inc: sc-81940), G3BP2 (Bethyl Laboratories, Inc: A302-040A), TIA-1 (Santa Cruz Biotechnology, Inc: sc-1751), TIAR (Santa Cruz Biotechnology, Inc: sc-1749), eIF3b (Santa Cruz Biotechnology, Inc: sc-16377), eIF4G (Santa Cruz Biotechnology, Inc: sc-11373), eIF4E-BP1 (Cell Signaling Technology: 9452S), PABP (Santa Cruz Biotechnology, Inc: sc-32318), Nucleolin (Santa Cruz Biotechnology, Inc: sc-9893), eIF4E (Santa Cruz Biotechnology, Inc: sc-9976), HuR (Santa Cruz Biotechnology, Inc: sc-5261), Fxr2 (Santa Cruz Biotechnology, Inc: sc32266), Rack1 (Santa Cruz Biotechnology, Inc: sc-17754), RPS23 (Santa Cruz Biotechnology, Inc: sc-100837), Twist1 (Bethyl Laboratories, Inc: A301-394A), Snail1 (Origene: TA500416), Zeb1 (Bethyl Laboratories, Inc: A301-921A), Puromycin (EMD Milipore: 12D10), GFP (Applied Biological Materials: G160), β-actin (Proteintech Group: 66009-1).
Lyons S.M., Achorn C., Kedersha N.L., Anderson P.J, & Ivanov P. (2016). YB-1 regulates tiRNA-induced Stress Granule formation but not translational repression. Nucleic Acids Research, 44(14), 6949-6960.
+ Open protocol
+ Expand
10

Measuring eIF4F Cap Complex Formation

Cited 1 time
Check if the same lab product or an alternative is used in the 5 most similar protocols
eIF4F cap complex formation was measured with m7-GTP batch chromatography as described (26 (link), 27 (link)). Briefly, cells were lysed in 20 mmol/L Tris-HCl (pH 7.4), 150 mmol/L NaCl, 1 mmol/L EDTA, 1 mmol/L EGTA, 1 mmol/L β-glycerophosphate, 1 mmol/L sodium orthovanadate, 1% Triton X-100, 0.2 mmol/L PMSF, 1× phosphatase inhibitor cocktails II and III (Sigma-Aldrich), and 1× HALT protease inhibitor cocktail (Thermo Scientific) for 15 minutes on ice. Lysate (400 μg) was incubated with 200μL of Immobilized γ-Aminophenyl-m7-GTP Agarose (Jena Bioscience) overnight at 4°C. Beads were washed 3 times with lysis buffer; bound protein was eluted, denatured, and then separated with SDS-PAGE followed by immunoblotting for eIF4G (Santa Cruz), 4E-BP1, and eIF4E (Cell Signaling).
Wahba A., Rath B.H., Bisht K., Camphausen K, & Tofilon P.J. (2016). Polysome profiling links translational control to the radioresponse of glioblastoma stem-like cells. Cancer research, 76(10), 3078-3087.
+ Open protocol
+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required

Sign up now

Revolutionizing how scientists
search and build protocols!