DNA Sequencing

Sanger Method

Answer the following questions after completing the activity DNA Sequencing: The Sanger Method.   You may find it helpful to watch the animation of the Sanger Method of DNA sequencing as a quick review.

  1. What are the four major steps of a DNA sequencing reaction?
  2. How many reaction tubes are required to sequence a small portion of DNA?
  3. List the components of the reaction that are the same in each microtube.  What is the purpose of each component?
  4. Name the reaction component that differs in each tube.
  5. What is the common name/phrase used to describe a dideoxyribonucleotide?
  6. Describe the process of DNA sequencing via the Sanger method.
  7. How many wells are required when loading a gel?
  8. After gel electrophoresis has been completed, does one determine the sequence beginning at the anode or cathode end of the gel?  Why?
  9. What is the name given to the “x-ray” of the gel?
  10. Is the sequence represented by the gel identical to the template DNA?

Automated Sequencing

In preparation for answering the following questions, watch animated tutorials of automated sequencing from Wiley and PBS and consider the electropherogram activity conducted in class.

  1. How many reaction tubes are required to perform automated DNA sequencing?  Explain why this differs from manual (Sanger) sequencing.
  2. What type of electrophoresis is used in automated sequencing?
  3. What detects the presence of a fluorescent dye?
  4. What is the name given to the “graph” produced in automated sequencing?  What does it look like?

General Questions

  1. Consider the activity “Assembling a DNA Sequence.”  How does one assemble a large sequence from several smaller fragments?
  2. What is a contig?
  3. What is depicted in the picture?   
  4. What is this?  How was it obtained?

electropherogram.jpg

(graphic obtained from Sierra College)

Autorads and electropherograms purchased from Edvotek.

edvotek-logo.jpg

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