Southern Blotting
Southern Blotting - Related Videos
02:57
Southern Blot
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2021
Agarose gel electrophoresis is very useful in separating DNA fragments by size. Running a DNA ladder containing fragments of the known length alongside the sample helps determine the approximate length of the sample DNA fragments. However, additional steps are needed to verify the sequence identity of the sample DNA fragments. Denatured DNA fragments must be transferred onto a carrier membrane from the gel to make it accessible to a probe - a small ssDNA fragment complementary to the target DNA...
05:42
Southern Blotting to Detect Homologous Recombination in the Mouse Embryonic Stem Cell Genome
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2025
In this video, we describe the Southern blotting technique to identify homologous recombination events in the genome of mouse embryonic stem cells using radiolabeled probes. DNA regions containing homologous recombination events appear as distinct bands when the membrane containing the probe-bound DNA is exposed to X-rays.
08:01
Identification of Homologous Recombination Events in Mouse Embryonic Stem Cells Using Southern Blotting and Polymerase Chain Reaction
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Cited by 1 •
2018
Here, we present a detailed protocol for identifying homologous recombination events that occurred in mouse embryonic stem cells using Southern blotting and/or PCR. This method is exemplified by the generation of nonmuscle myosin II genetic replacement mouse models using traditional embryonic stem cell-based homologous recombination-mediated targeting technology.
08:48
The Western Blot
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2023
Western Blotting is used to identify the presence of specific proteins in electrophoretically separated samples. Following separation by a technique known as sodium dodecyl sulfate polyacrylamide gel electrophoresis, or SDS-PAGE, western transfer is used to move proteins from a polyacrylamide gel onto a piece of membrane which traps the proteins in their respective locations. Next, the membranes are probed with antibodies in a process called immunboblotting. Immunoblotting uses...
01:15
Western Blotting
0 Views •
2023
Western blotting is an analytical technique for protein identification. It has various applications in immunology and medicine, including detecting diseases like bovine spongiform encephalopathy, mad cow disease, and human and feline immunodeficiency virus from biological samples. The technique begins with separating proteins from the sample using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), followed by protein transfer, immunoblotting, and finally, protein detection.
FAQs
Restriction enzymes determine which DNA fragments become available for downstream analysis. By cutting genomic DNA at defined recognition sites, they generate a reproducible fragment pattern. That pattern matters because a target sequence can be identified in a fragment whose length and neighboring arrangement differ after a structural change. Comparing patterns therefore helps reveal altered gene organization.
The labeled probe supplies the sequence-level specificity that distinguishes a target fragment from the many other DNA fragments on the membrane. Because it is complementary to the selected sequence, hybridization identifies fragments carrying that sequence. The label then makes the bound probe detectable, allowing researchers to connect a molecular signal with a particular genomic fragment.
Southern blotting is especially informative when the question concerns a DNA fragment’s size or genomic arrangement, not merely whether a sequence can be detected. Although amplification methods and sequencing are often faster, the blot provides a fragment-pattern view that supports mapping, structural analysis, and validation of engineered DNA.
An effective workflow preserves the link between digestion, separation, transfer, and probe detection. After restriction enzymes cut the sample, gel electrophoresis separates fragments by size. Denaturation prepares them for probe binding, and transfer to a membrane fixes the separated pattern in a format suitable for hybridization. Each stage retains information needed for interpretation.
Fragment size and band arrangement provide complementary evidence. A detected target in an expected fragment supports its presence, while a changed fragment pattern can indicate altered DNA structure or organization. Multiple fragments can also support gene mapping by showing where the selected sequence occurs relative to restriction sites. These interpretations make the method useful beyond simple detection.
In biology, Southern blotting can support gene mapping, genotyping, and examination of structural changes in genomic DNA. It also helps validate cloned or engineered DNA by checking whether the expected target sequence and fragment organization are present. These applications make the technique useful when physical DNA arrangement matters, even though faster methods may be preferred for some analyses.