Interpret the provided gel photo: Did the restriction enzymes appear to successf

Question

Interpret the provided gel photo:
Did the restriction enzymes appear to successfully cut the plasmid DNA?
• Look for differences between the undigested and digested samples.
• Do the observed bands match the predictions (based on knowledge of the plasmids)?

Figure 5. Agarose gel electrophoresis of plasmid DNA before and after digestion 1 2 3 4 5 6 7 8 9 10 11 reactions. Electrophoresis image of 1% agarose gel stained with GelRed and visualised under UV light. Sample preparation and gel electrophoresis was performed by Garry Penney Lane 1 Markers (SPP1 EcoRI digest) Lane 2 pK184hlyA undigested Lane 3 pK184hlyA digested with EcoRI Lane 4 pK184hlyA digested with Pstl Lane 5 pK184hlyA Lane 6 Lane 7 digested with EcoRI and Pstl Markers (SPP1 EcoRI digest) pBluescript digested with EcoRI and Pstl Lane 8 pBluescript digested with EcORI Lane 9 pBluescript digested with Pstl Lane 10 pBluescript undigested Lane 11 Markers (SPP1 EcoRI digest) Copy of Figure 2. SPP1 EcoRI size markers after electrophoresis Wells through 1% agarose gel When assigning sizes of marker bands on 1% agarose gels like this, determine the bands from the top (i.e. largest bands); the smallest fragments of DNA in the marker lanes have already run off the gel in Figure 5 SPP1 EcoRI size markers consist of the DNA of SPP1 phage digested with EcoRI restriction enzyme. Size (kb) markers are used in gel electrophoresis to estimate the sizes of DNA fragments of unknown length. 851 7.35 6.11 4.84 3.59 281 1.95 1.86 1.51 1.39 1.16 0.98 0.72 0.48 0.36

Explanation / Answer

Answer. Yes in case of pK184hIyA restriction enzyme cut the plasmid DNA as in lane 2 undigested which travels farthest as compared to lane 3 and 4. It means plasmid have one restriction site for EcorI and one restriction site for PstI , therefore two band were seen in lane. That means PstI and EcorI cut at different site and creates two different size of cut plasmid.

In case of pBluescript plasmid in lane 7 digested with EcorI and PstI show only one band but it is darker as compared to lane 8 and 9, the conclusion is that the PstI and EcorI cut plasmid at same distance which gives only one band while compared to lane 10 undigested pBluescript plasmid it travel farther that means it is a circular plasmid. So it travel farther.

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