![]() ![]() Importantly, the ES cells being used already had a modification that allowed researchers to easily insert a gene of interest. įigure 2 above shows the results of a two Southern blots, the first from embryonic stem cells that the authors were attempting to genetically modify, and the second from the resulting genetically modified mice. (C) Southern blot analysis of SpeI-digested genomic DNA from mice of the indicated genotypes. Clones D4 and D5 exhibit proper targeting of the Col1A locus and an unaffected wild-type allele. Relevant section of caption for published figure reads: “ (B) Southern blot analysis of SpeI-digested genomic DNA from six PHGDHtetO-targeted ES cells. “Southern Blot Illustration” by National Human Genome Research Institute. The film will then show bands at the location where the DNA sequence of interest was found on the filter, and ultimately, on the agarose gel.įigure 1. If the probe is radioactive, this is done by exposing an X-ray film to the filter. This leave invisible radioactive bands on the filter where the DNA sequence of interest is located. Once the probe has hybridized with the DNA on the filter, excess probe is washed away. The probe is a nucleic acid sequence that is complementary to the genomic DNA of interest. Usually, the probe is labeled with 32P, which is radioactive. Once the DNA is transferred to the filter, the filter is incubated with a labeled probe. Then, as shown in Figure 1 below, the DNA from the agarose gel is transferred onto a nitrocellulose filter by allowing a buffer to run through the gel, then the membrane, into a stack of dry blotting paper. The resulting DNA fragments are separated by gel electrophoresis (insert link). Once genomic DNA is obtained, it is usually digested using one or more restriction enzymes. Samples needed: Genomic DNA from cells or sample of interestĬontrols: Controls should be performed to test a new probe but are often not shown in a publication. Often it is used to determine if a planned genetic modification has been successfully accomplished. ![]() A Southern blot shows the size of a piece of DNA with a particular sequence and is mostly used on digested genomic DNA. ![]()
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