Map-based Cloning
Map-based Cloning - Related Videos
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...
04:25
CRISPR-based Shuttle Cloning: A High-throughput Cloning Method
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2025
We describe a protocol for a high-throughput cloning method, CRISPR-based shuttle cloning (CRISPRshuttle cloning), which allows the transfer of DNA fragments of interest between vectors without the need for PCR amplification of the DNA fragments.
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.
06:28
Standardized Modular Assembly of Polycistronic Operons with Modular Cloning (MoClo) using the In-Cloning toolkit
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2025
This article presents a step-by-step protocol demonstrating how Modular Cloning (MoClo) can be adapted for the cloning of polycistronic operons.
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.
FAQs
Linkage analysis examines whether inheritance of a trait consistently occurs alongside particular molecular markers. A marker that tracks with the phenotype indicates that both lie near one another on the same chromosome, because nearby positions are less likely to be separated during recombination. This association provides the starting point for locating the responsible gene.
Recombination mapping uses differences in chromosome rearrangement among individuals to refine the trait-associated interval. Recombination events that separate a marker from the phenotype help establish which chromosome segments are unlikely to contain the causal gene. Repeatedly comparing these boundaries can reduce a broad linked region to a smaller segment containing fewer possible genes.
Chromosome walking progressively examines overlapping DNA fragments across the mapped interval. Each fragment extends the analysis from a known position toward adjacent genomic sequence, helping researchers build a continuous view of the region. This approach is especially useful when the initial marker association identifies a location but does not yet reveal which gene produces the trait.
Researchers examine the genes located within the refined interval by sequencing them and looking for mutations associated with the phenotype. They then consider functional evidence to determine whether a candidate plausibly accounts for the trait. Combining positional information with sequence and functional results strengthens the case for identifying the responsible gene rather than selecting a candidate based only on location.
A typical workflow begins by associating the phenotype with molecular markers through linkage analysis. Researchers then use recombination mapping to narrow the interval, examine overlapping DNA fragments through chromosome walking, and identify genes within the resulting region. Sequencing and functional testing of candidate genes provide evidence that connects a particular gene to the observed trait.
This strategy is useful when researchers need to uncover the genetic basis of an inherited trait, disease phenotype, or agriculturally important characteristic. By connecting a phenotype to a specific gene, the approach can reveal gene function and support genetic research. In agriculture, the resulting knowledge can also contribute to molecular breeding efforts focused on important characteristics.