Fluorescent Protein-Tagged Plasmids

The following plasmids were used as received from Addgene: mRuby-Golgi-7 (GalT; #55865), mRuby2-Rab5a-7 (#55911), mCherry-Rab7a-7 (#55127), mCherry-Rab11a-7 (#55124), pmCherry-2xFYVE (#140050). Venus-2xFYVE was made by replacing mCherry in pmCherry-2xFYVE with Venus using NheI and BsrGI. mRuby2-MOA was made by replacing Venus in Venus-MOA using NheI and BglII. mRuby2-PTP1b was made by replacing Venus in Venus-PTP1b using NheI and BsrGI. CMV-LgBit was made by amplifying LgBit from pBiT1.1-N (Promega) and ligating into pcDNA3.1 (+) using HindIII and XhoI. SNAPf-β₂AR, SNAPf-D2R, SNAPf-M3R and D2S-Nluc were kindly provided by Jonathan Javitch (Columbia University). Venus-Kras, Venus-PTP1b, Venus-MOA, Venus-rab5a, Venus-rab7a, Venus-rab11a and memGRKct-Venus were described previously (Hollins et al., 2009 ; Lan et al., 2012 (link)). All plasmids were verified by automated sequencing.

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Jang W., Senarath K., Lu S, & Lambert N.A. (2024). Visualization of endogenous G proteins on endosomes and other organelles. bioRxiv.

Publication Preprint 2024

pBiT1.1-N

Corresponding Organization : Augusta University

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Variable analysis

independent variables

Replacing Venus in Venus-MOA with mRuby2 using NheI and BglII
Replacing Venus in Venus-PTP1b with mRuby2 using NheI and BsrGI
Replacing mCherry in pmCherry-2xFYVE with Venus using NheI and BsrGI
Amplifying LgBit from pBiT1.1-N and ligating into pcDNA3.1 (+) using HindIII and XhoI

dependent variables

Not explicitly mentioned

control variables

All plasmids were received as is from Addgene, except for the modified versions

controls

No positive or negative controls were explicitly mentioned

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