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Wrap up MiniPrep. Analysis and conclusion. What does the gel Show? Why might there be 3 bands for a purified plasmid?. Might have a very light line of “nicked” Plasmid DNA that “Runs” at 2X its actual bp length. Plasmid “ripped” open!. Most of it “Runs” like a linear piece of

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Wrap up miniprep

Wrap up MiniPrep

Analysis and conclusion


Wrap up miniprep

What does the gel Show?

Why might there be 3 bands for a purified plasmid?

Might have a very light line of “nicked” Plasmid

DNA that “Runs” at 2X its actual bp length

Plasmid “ripped” open!

Most of it “Runs” like a linear piece of

DNA 0.5X the plasmids actual bp length

p. 2B-10


What does the gel show
What does the gel show?

  • Did you run a standard plasmid?

    • Can use to analyze the plasmid DNA concentration of your miniprep!

    • EX.

      If used concentration :

      Plasmid sample prepped with orange G?

      ()()=

      3.0 of S3 plasmid

      Miniprep+orange G a total of 12, loaded 10., there fore:

      ()(3.0 of S3 plasmid)=

      2.5 ngof S3 plasmid loaded into well

      How can this information be used???


Results
Results

  • Raw:

    • Figure(s) of Gel

      • Fully annotate! What should you put!

    • Direct observations

      • FYI: once you run a gel, we talk about lane #, not well #

  • Analysis:

    • What mass of DNA was loaded into Standard Lane 1?

      • Show all calculations with descriptive anotations!

    • By visual comparison, Is it possible to calculate the mass of DNA loaded into sample in __ lane?

    • Is it possible to calculate the concentration of plasmid DNA in the Yellow MCT in the freezer?


Is it possible to calculate the concentration of plasmid dna in the yellow mct in the freezer
Is it possible to calculate the concentration of plasmid DNA in the Yellow MCT in the freezer?

  • How much of the plasmid solution was used when prepping w/ orange G?

  • What fraction of the plasmid/orange G solution was loaded into the well?

  • How many nanograms of plasmid was in lane __?

  • Concentration units should be in


Conclusion
Conclusion in the Yellow MCT in the freezer?

  • Is there evidence that plasmid was purified from the transformed E.coli HB101?

    • What is the evidence?????

    • What can be concluded from the presence or absence of band(s) in the lane???