Gw1 mito phred

Manufactured by Addgene

The GW1-Mito-pHRed is a fluorescent protein that can be used to monitor pH changes in the mitochondria of live cells. It is a genetically-encoded pH indicator that specifically targets the mitochondria, allowing researchers to visualize and quantify pH dynamics within this cellular organelle.

Automatically generated - may contain errors

Lab products found in correlation

Pcdna3 flag rheb, by Addgene (1 mentions) Memerald mito 7, by Addgene (1 mentions) Shbnip3l, by Addgene (1 mentions) Mcherry mito 7, by Addgene (1 mentions) Plko 1 puro, by Addgene (1 mentions) Q5 site directed mutagenesis kit, by New England Biolabs (1 mentions) Pultra hot, by Addgene (1 mentions) Pegfp lc3, by Addgene (1 mentions) Jetprime reagent, by Polyplus Transfection (1 mentions)

4 protocols using gw1 mito phred

Plasmids for BNIP3L Protein Characterization

Cited 5 times

Check if the same lab product or an alternative is used in the 5 most similar protocols

MYC-tagged BNIP3L (Myc-Nix), BNIP3L-CYB5 (Nix-CytoB5), BNIP3L-MaoB (Nix-MaoB), BNIP3L-ActA (Nix-ActA), BNIP3L-ΔTM (Nix-delta TM) plasmids (Addgene, 100,795, 100,756, 100,757, 100,758, and 100,755; deposited by Joseph Gordon) were described previously [31 (link)]. The MYC-tagged BNIP3L^S212A and BNIP3L^S212D were generated using a Q5 site directed mutagenesis kit (New England Biolabs, E0554S). The lentiviral shBNIP3L (Addgene, 100,770; deposited by Joseph Gordon) was generated by ligating oligonucleotides containing the targeting sequence 5ʹ-CAGTTCCTGGGTGGAGCTA-3ʹ into pLKO.1-puro (Addgene, 8453; deposited by Bob Weinberg). The mitochondrial (CMV-mitoCAR-GECO1) and endoplasmic reticulum (CMV-ER-LAR-GECO1) targeted calcium biosensors were gifts from Robert Campbell (Addgene, 46,022 and 61,244; deposited by Robert Campbell) [32 (link),33 (link)]. The shBNIP3L (17,469; deposited by Wafik El-Deiry) [54 (link)], mEmerald-Mito-7 and mCherry-Mito-7 (54,160 and 55,102; deposited by Michael Davidson) [55 ,56 (link)], pPHT-PRKA (60,936; deposited by Anne Marie Quinn) [57 (link)], DAGR (14,865; deposited by Alexandra Newton) [37 (link)], GW1-Mito-pHRed (31,474; deposited by Gary Yellen) [36 (link)] and pcDNA3-FLAG-RHEB (19,996; deposited by Fuyuhiko Tamanoi) [58 (link)] plasmids were purchased from Addgene.

da Silva Rosa S.C., Martens M.D., Field J.T., Nguyen L., Kereliuk S.M., Hai Y., Chapman D., Diehl-Jones W., Aliani M., West A.R., Thliveris J., Ghavami S., Rampitsch C., Dolinsky V.W, & Gordon J.W. (2020). BNIP3L/Nix-induced mitochondrial fission, mitophagy, and impaired myocyte glucose uptake are abrogated by PRKA/PKA phosphorylation. Autophagy, 17(9), 2257-2272.

+ Open protocol

+ Expand

Investigating Mitochondrial pH Dynamics

Check if the same lab product or an alternative is used in the 5 most similar protocols

A mito-pH indicator was used to investigate mitochondrial pH fluctuations that may interact with our biosensor sensitivity. The pH indicator used was GW1-Mito-pHRed, a gift from Gary Yellen via Addgene (Addgene plasmid #31474, accessed 6 June 2022). A control experiment in normoxia was run post-transfection of HEK293T cells with mito-pH tracker for 2 h, and then an additional run was run in hypoxic conditions for the same time (Figure S3).

White D I.I.I., Lauterboeck L., Mobasheran P., Kitaguchi T., Chaanine A.H, & Yang Q. (2023). Real-Time Visualization of Cytosolic and Mitochondrial ATP Dynamics in Response to Metabolic Stress in Cultured Cells. Cells, 12(5), 695.

+ Open protocol

+ Expand

Ratiometric pH Sensors Expression

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

SypHer mt (Addgene #48251), GW1-Mito-pHRed (Addgene #31474), and pUltra-hot (Addgene #24130) were gifts from Nicolas Demaurex, Gerry Yellen, and Malcolm Moore, respectively. SypHer mt allows the expression of a pH-sensitive ratiometric cpYFP derivative that contains two mitochondrial matrix localization sequences at its N-terminus.²⁸ (link)^,²⁹ (link) GW1-Mito-pHRed encodes for a pH-sensitive ratiometric m-Keima derivative that contains four cytochrome C oxidase subunit VIII (Cox 8) tags to target it to the mitochondria.³⁰ (link) We cloned the pH-sensors into the lentiviral expression vector pUltra-hot to produce viruses for an efficient, moderate, and stable expression of the pH indicators in our target cells. For cloning procedure, please see SI.

Schaefer P.M., Hilpert D., Niederschweiberer M., Neuhauser L., Kalinina S., Calzia E., Rueck A., von Einem B, & von Arnim C.A. (2017). Mitochondrial matrix pH as a decisive factor in neurometabolic imaging. Neurophotonics, 4(4), 045004.

+ Open protocol

+ Expand

Transfection and Mitochondrial Imaging

Check if the same lab product or an alternative is used in the 5 most similar protocols

H9c2s were cultured and transfected with plasmids using JetPRIME Reagent (PolyPlus). After incubation with the transfection media for 6 hours, the media was replaced with normal culture media and incubated overnight at 37°C. The next day, the cells were treated with DOX and imaged 24 hours later. GW1-Mito-pHRed (Addgene 31474) was a gift from Gary Yellen (34) (link); and pEGFP-LC3 (Addgene, 24920) was a gift from Toren Finkel (35) .

Kawalec P., Martens M.D., Field J.T., Mughal W., Caymo A.M., Chapman D., Xiang B., Ghavami S., Dolinsky V.W, & Gordon J.W. (2022). Differential impact of doxorubicin dose on cell death and autophagy pathways during acute cardiotoxicity. Toxicology and applied pharmacology, 453.

+ 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?

Revolutionizing how scientists
search and build protocols!