Plasmid Cloning
Plasmid 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...
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.
08:30
Yeast Transformation and Cloning
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2023
S. cerevisiae are unicellular eukaryotes that are a commonly-used model organism in biological research. In the course of their work, yeast researchers rely upon the fundamental technique of transformation (the uptake of foreign DNA by the cell) to control gene expression, induce genetic deletions, express recombinant proteins, and label subcellular structures. This video provides an overview of how and why yeast transformation is carried out in the lab. The important features of yeast...
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...
07:11
Homemade Site Directed Mutagenesis of Whole Plasmids
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Cited by 91 •
2009
Site directed mutagenesis of whole plasmids is a simple way to create slightly different variations of an original plasmid. Here we demonstrate an easy and cost effective way to introduce base substitutions into a plasmid using standard reagents.
FAQs
The insert supplies the DNA sequence selected for copying, analysis, or expression, whereas the plasmid vector provides a circular, self-replicating context that can be maintained in host cells. Their combination creates a recombinant plasmid, allowing researchers to study the sequence within a biological system rather than only as isolated DNA.
Restriction enzymes and DNA ligase support complementary stages of construct assembly. Restriction enzymes prepare the plasmid and insert for joining, while DNA ligase connects them into a continuous recombinant DNA molecule. When a compatible assembly method is used instead, it provides an alternative way to combine the same essential components.
Selection, screening, and sequencing provide progressively stronger evidence for the intended construct. Antibiotic selection identifies cells carrying the vector, but it does not by itself establish that the insert is correct. Colony screening evaluates candidate colonies, and DNA sequencing confirms the intended DNA arrangement for downstream biological work.
A typical workflow begins by combining a DNA insert with a plasmid vector through restriction-enzyme and ligase treatment or a compatible assembly method. The resulting recombinant plasmid is introduced into bacteria by transformation. Antibiotic selection identifies vector-containing cells, followed by colony screening and DNA sequencing to confirm suitable constructs.
A confirmed construct can support several types of investigation. Researchers may copy the selected DNA sequence, analyze it, or examine its expression in host cells. Because colony screening and sequencing verify the intended construct, subsequent experiments can connect observed biological results more confidently to the inserted sequence.
Plasmid cloning supports gene function studies, protein production, genetic engineering, and the development of research tools. In biology, these applications make it possible to place selected DNA sequences into recombinant plasmids and evaluate their behavior or use. The method therefore connects DNA construction with functional and applied research outcomes.