Dna Ligation Reaction
Dna Ligation Reaction - Related Videos
07:34
DNA Ligation Reactions
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2023
In molecular biology, ligation refers to the joining of two DNA fragments through the formation of a phosphodiester bond. An enzyme known as a ligase catalyzes the ligation reaction. In the cell, ligases repair single and double strand breaks that occur during DNA replication. In the laboratory, DNA ligase is used during molecular cloning to join DNA fragments of inserts with vectors – carrier DNA molecules that will replicate target fragments in host organisms. This video provides an...
13:10
Detection and Visualization of DNA Damage-induced Protein Complexes in Suspension Cell Cultures Using the Proximity Ligation Assay
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Cited by 3 •
2017
Here, it is demonstrated how the in situ Proximity Ligation Assay (PLA) can be used to detect and visualize the direct protein-protein interactions between ATM and p53 in suspension cell cultures exposed to genotoxic stress.
03:02
Microsphere Polymerase Chain Reaction to Amplify Single-Stranded DNA
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2025
This video describes the technique of amplifying single-stranded DNA using oligonucleotide-coated polymer microspheres in a single-tube reaction. This process yields pure single-stranded DNA, which can be further used for DNA sequencing and microarray analysis.
09:31
Amplification, Next-generation Sequencing, and Genomic DNA Mapping of Retroviral Integration Sites
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Cited by 56 •
2016
We describe a protocol for amplifying retroviral integration sites from the genomic DNA of infected cells, sequencing the amplified virus-host junctions, and then mapping these sequences to a reference genome. We also describe techniques to quantify the distribution of integration sites relative to various genomic annotations using BEDTools.
04:28
Multiplex Droplet Polymerase Chain Reaction to Detect DNA Methylation in Leukocytes
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2025
This video demonstrates the multiplex droplet polymerase chain reaction technique for the simultaneous quantification of methylated DNA regions in different white blood cell populations as an early biomarker for clinical diagnosis and prognosis.
FAQs
Cohesive ends generally improve joining efficiency because complementary single-stranded sequences can align adjacent DNA fragments before the enzyme seals them. Blunt-ended fragments lack this complementary overhang, so they can still be joined, but only under suitable conditions. This distinction helps researchers choose compatible fragment designs when assembling defined sequences.
DNA ligase requires an energy source to drive bond formation, with ATP or NAD+ serving this role in the reaction. The cofactor choice is therefore a mechanistic condition rather than a structural feature of the DNA fragments. Recognizing this requirement helps explain why ligation performance depends on using conditions that support ligase activity.
The two reactive DNA termini must be positioned on adjacent strands: a 3′ hydroxyl group and a 5′ phosphate group provide the chemical partners for bond formation. If the required groups are not aligned at the junction, the enzyme cannot efficiently create the continuous backbone needed for a stable joined product.
A practical ligation design starts by identifying whether the DNA fragments carry complementary cohesive ends or blunt ends, then selecting reaction conditions compatible with the ligase and its energy source. The outcome is assessed by whether the intended fragments become joined. This planning is especially important when constructing defined recombinant DNA molecules.
Cloning and plasmid construction use ligation to combine selected DNA fragments into a defined recombinant arrangement. This allows researchers to build molecules that can subsequently support analysis or expression of genetic sequences. The method is particularly useful when the desired construct depends on joining specific fragments rather than examining them as separate pieces.
DNA library preparation applies ligation to assemble collections of DNA molecules for later analysis, while molecular diagnostics uses the same joining capability to construct or examine defined genetic arrangements. These applications show that the reaction is not limited to one type of experiment: its value comes from controlled fragment joining across workflows with different analytical goals.