Mitochondrial Ligase 1 Localization

Rabbit anti-FUS (Cat# A300–302 A) and anti-Lig1 (A301-136A) antibodies were procured from Bethyl Laboratories, Inc. Mouse anti-FLAG antibody (A8592) was obtained from Sigma-Aldrich, and mouse anti-Lig3 antibody (Cat# ab587) was purchased from Abcam. Mouse anti-Tom20 (SC-17764), anti-HSP60 (SC-13115) and anti-PCNA (SC-56) antibodies were procured from Santa Cruz Biotechnology. Rabbit anti-CYTB (Cat# 55090-1-AP), COX1 (Cat# 13393-1-AP) and ND4 (Cat# 26736-1-AP) procured from Protein tech. Fluorescent secondary antibodies, Alexa Fluor 488 anti-mouse (Cat# A28175), and Texas Red anti-rabbit antibody (Cat# T-2767) were obtained from Life Technologies. The antibodies were diluted at 1:1000 for western blotting, 1:500 for immunofluorescence, and 1:100 for PLA.
The human Lig1 coding sequence was re-cloned into pCDNA3.1 plasmid as an N-terminal FLAG and COX8 gene mitochondrial targeting sequence containing construct (FLAG-MTS-Lig1)^{56 (link)}.

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Kodavati M., Wang H., Guo W., Mitra J., Hegde P.M., Provasek V., Rao V.H., Vedula I., Zhang A., Mitra S., Tomkinson A.E., Hamilton D.J., Van Den Bosch L, & Hegde M.L. (2024). FUS unveiled in mitochondrial DNA repair and targeted ligase-1 expression rescues repair-defects in FUS-linked motor neuron disease. Nature Communications, 15, 2156.

Publication 2024

Rabbit anti-FUS anti-Lig1 (A301-136A) Mouse anti-FLAG antibody (A8592) mouse anti-Lig3 antibody anti-PCNA (SC-56) Alexa Fluor 488 anti-mouse Texas Red anti-rabbit antibody

Corresponding Organization : Cornell University

Other organizations : Houston Methodist, VIB-KU Leuven Center for Brain & Disease Research, BioEnergetics (United States), New Mexico Cancer Center, University of New Mexico

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Variable analysis

independent variables

Overexpression of FLAG-MTS-Lig1 construct

dependent variables

Protein expression levels of Lig1, FUS, Lig3, Tom20, HSP60, PCNA, CYTB, COX1, and ND4

control variables

Antibody dilutions for western blotting (1:1000), immunofluorescence (1:500), and PLA (1:100)

controls

Positive controls: Not specified
Negative controls: Not specified

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