Marcm Clones
Marcm Clones - Related Videos
05:48
A Behavioral Assay for Mechanosensation of MARCM-based Clones in Drosophila melanogaster
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Cited by 1 •
2015
In order to identify novel mutations affecting mechanosensation, we designed an assay that measures the behavioral response to tactile stimulation of fly bristles in mutant clones generated by the MARCM method. The combination of techniques allows for the identification of mechanosensitive mutations that would otherwise be lethal.
06:30
Cell Lineage Analyses and Gene Function Studies Using Twin-spot MARCM
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Cited by 2 •
2017
Here, we present a protocol for a mosaic labeling technique that permits the visualization of neurons derived from a common progenitor cell in two distinct colors. This facilitates neural lineage analysis with the capability of birth-dating individual neurons and studying gene function in the same neurons of different individuals.
09:55
Molecular Cloning
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2023
Molecular cloning is a set of methods, which are used to insert recombinant DNA into a vector - a carrier of DNA molecules that will replicate recombinant DNA fragments in host organisms. The DNA fragment, which may be a gene, can be isolated from a prokaryotic or eukaryotic specimen. Following isolation of the fragment of interest, or insert, both the vector and insert must be cut with restriction enzymes and purified. The purified pieces are joined together though a technique called...
01:27
Reproductive Cloning
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2019
Reproductive cloning is the process of producing a genetically identical copy—a clone—of an entire organism. While clones can be produced by splitting an early embryo—similar to what happens naturally with identical twins—cloning of adult animals is usually done by a process called somatic cell nuclear transfer (SCNT). Somatic Cell Nuclear Transfer In SCNT, an egg cell is taken from an animal and its nucleus is removed, creating an enucleated egg. Then a somatic cell—any cell that is not a sex...
05:21
Culturing and Harvesting of HSPC Clones: A Method to Obtain the Singly-Sorted HSPC Clones
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2023
This video describes the protocol for isolating HSPCs from the bone marrow and sorting them using a fluorescent activated cell sorting technique, which is a method to obtain singly-sorted HSPCs. We culture these sorted cells to obtain HSPC clones.
FAQs
During mitosis, FLP-mediated recombination at matching FRT sites produces daughter-cell lineages with different genetic combinations. This can make a clone homozygous for a mutation while surrounding tissue remains genetically normal or distinct. The resulting contrast lets investigators attribute changes in a cell's development or morphology to its genotype rather than to a mutation throughout the animal.
GAL80 normally represses GAL4-driven transcription, so its loss within recombined cells releases expression of a fluorescent or other cell marker. This creates a genetic link between clone formation and visualization: the relevant cells become labeled specifically. Researchers can therefore identify mutant cells while examining their position, morphology, and relationships within neural tissue.
Because MARCM clones restrict the genetic change to selected cells, they help separate effects originating in labeled cells from consequences of a mutation throughout the organism. In Drosophila neuroscience, this is useful when broad mutations could alter surrounding tissue or development. Comparing clone phenotypes with genetically normal neighbors strengthens links between genotype and neuronal structure or function.
MARCM clones support analysis of neuronal development, dendritic and axonal morphology, synaptic connectivity, and circuit function. These readouts span how a neuron develops, builds processes and synaptic relationships, and participates in a functional circuit. Using several readouts can reveal whether a gene affects structure, connectivity, function, or more than one level.
A MARCM design brings together FLP/FRT-mediated recombination, GAL80-mediated repression, GAL4-driven marker expression, and a fluorescent or other cell marker. Their roles are complementary: recombination creates genetically distinct cells, GAL80 loss permits labeling, and the marker makes the clone traceable. Together, these components allow genetic identity and anatomical observations to be analyzed in neural tissue.
At single-cell resolution, investigators can relate a clone's genotype to neuronal structure, synaptic connectivity, and circuit function, then consider consequences for behavior. This makes MARCM useful when the question concerns how particular neural cells contribute to a larger circuit. Genetically normal surrounding tissue also helps distinguish clone-associated effects from broad organism-wide effects.