specific-labeling-mitochondrial-nucleoids-for-time-lapse-structured

Specific Labeling of Mitochondrial Nucleoids for Time-lapse Structured Illumination Microscopy

Mitochondrial nucleoids are compact particles formed by mitochondrial DNA molecules coated with proteins. Mitochondrial DNA encodes tRNAs, rRNAs, and ...

Max Planck Institute of Immunobiology and Epigenetics

Mitochondrial DNA molecules coated with proteins form compact particles called mitochondrial nucleoids. They are redistributed within mitochondrial network undergoing morphological changes. The straightforward technique to characterize nucleoids' motions is fluorescence microscopy. Mitochondrial nucleoids are commonly labelled with fluorescent protein tags, which is not always feasible and was reported to cause artifacts. Organic DNA-binding dyes are free of these drawbacks, but they lack specificity to mitochondrial DNA. Here, considering physico-chemical properties of such dyes, we achieved preferential live-cell labelling of mitochondrial nucleoids by a nucleic acid staining dye SYBR Gold. It enabled time-lapse imaging of mitochondrial nucleoids by structured illumination microscopy and quantification of their motions.

SYBR Gold dye enables preferential labelling of mitochondrial nucleoids and their time-lapse imaging by structured illumination microscopy.

Conventional fragments of fluorescent proteins used in bimolecular fluorescence complementation technique (BiFC), form light-emitting species only when they are kept in close proximity by interacting proteins of interest. By contrast, certain fluorescent protein fragments complement spontaneously, namely those corresponding to the 1st to 10th beta-strands (GFP1-10) and the 11th beta-strand of superfolder GFP (GFP11). They were designed as folding reporters for high throughput expression and structure biology. Besides, for light microscopy, self-associating fluorescent protein fragments constitute a valuable and sometimes unique tool. The GFP11 tag is very advantageous when a full-length fluorescent protein cannot be fused to a protein of interest, namely for live imaging of certain pathogens. Self-associating GFP fragments enable live labelling of specific synapses, visualization of proteins topology and their exposure to particular subcellular compartments. Present review aims to attract attention of scientific community to these tools and to inspire their further development and applications.

Sergiy V. Avilov

Imaging Facility

Max Planck Institute of Immunobiology and Epigenetics

Specific Labeling of Mitochondrial Nucleoids for Time-lapse Structured Illumination Microscopy

Sergiy V. Avilov

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Imaging Facility

Max Planck Institute of Immunobiology and Epigenetics

Specific Labeling of Mitochondrial Nucleoids for Time-lapse Structured Illumination Microscopy

Imaging Facility