Analyzing Gels

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Analyzing Gels

• Biotech 2

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Go online and find the answer to these questions

• What is a plasmid?
• What is pBR322 and what is its base pair (bp)
  size?
• What are the cut standards (base pair size) of
  Lambda DNA with HIND III? (PIC)

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What type of DNA can we use?

• Salmon Sperm
• Animal
• E.Coli
• Bacteria
• Yeast
• Eukaryotic
• Lambda
• Virus

• Lambda HIND III
• Virus DNA cut with an enzyme called HIND III
• pAMP
• Plasmid that makes an enzyme to break down
  ampicilin
• pBR322
• Most commonly used E.coli cloning plasmid

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What are plasmids?

• Small pieces of DNA
• Origin of Replication
• Antibiotic Resistance
• Promoter
• Gene of Interest

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What does a band (line) mean on a gel?

• Each band is a certain amount of base pairs (bp).
  
• With large number of base pairs at the top
• Smaller number base pairs at bottom

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How do we tell how big a piece of DNA is?

• Standards - Which one did we use?

• Lambda cut with HIND III - What are the sizes?

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Lambda

• What is your estimate of Lambda?
• Why is Lambda one line?

On the 0.8 agarose gel DNA gets hung up at
25,000 bp
48,000 bp
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Salmon Sperm

• What do you think the size of SS is?
• Why do you think it smears?

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E.coli

• What do you estimate the size of E.coli is?
• Why do you think it is different from Salmon
  Sperm?
• Why is one E.coli lighter than the other?

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pBR322
4361 bp
Supercoiled
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Yeast

• What is the size of Yeast?

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pUC19
2686 bp
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pAMP

• Which band on pAMP do we want to read?
• What do you think the size of pAMP is?

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Mistakes

• Look at the edges?
• What do you notice about the DNA?

HIGH VOLTAGE
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Mistakes

• Where is the DNA?

Poor Pipetting
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Mistakes

• What do you notice about the Salmon Sperm?

High concentration of DNA. Spills into other
wells.
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Mistakes

• What are some things you notice?

Agarose not fully melted, due to wavy lines.
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Mistakes

• Why is this incorrect?

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Reflection

• Label your DNA printout and analyze the results.

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References

• http//www.brunel.ac.uk/depts/bio/project/genome/m
  oltec/vectors/gifs/puc19.gif
• http//www.mun.ca/biochem/courses/4103/figures/Sny
  der-Champness/F4-1.jpg
• http//occawlonline.pearsoned.com/bookbind/pubbook
  s/bc_mcampbell_genomics_1/medialib/method/inducepr
  omoter.gif
• http//www.dnalc.org/images/pamp.jpg