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DNA Analysis using Gel Electrophoresis

DNA Analysis using Gel Electrophoresis. DNA Analysis. DNA can be analyzed in several different ways depending on the information that you want to find out DNA sequencing will tell you the letters in the sequence (Ex: AGTGGATATG . . .)

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DNA Analysis using Gel Electrophoresis

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  1. DNA Analysis using Gel Electrophoresis

  2. DNA Analysis • DNA can be analyzed in several different ways depending on the information that you want to find out • DNA sequencing will tell you the letters in the sequence (Ex: AGTGGATATG . . .) • Gel electrophoresis will look for specific sequences that can indicate the presence of specific genes

  3. DNA sequencing Gel electrophoresis

  4. Gel electrophoresis • Works by separating DNA segments (aka fragments) by placing them in a gel substance and applying an electrical current. • DNA fragments are sorted according to size

  5. How do we get these fragments? • To cut DNA in specific places, we use restriction enzymes (called restriction endonucleases) • Restriction enzymes cut DNA at specific sequences (like CCCTTT, they might cut between the last C and first T) • If you have that sequence in your DNA it will be cut, if not then it won’t. This results in different people having different DNA fragments. • http://www.dnalc.org/ddnalc/resources/restriction.html

  6. Why are we cutting the DNA? • We cut the DNA to see if the person has a specific sequence of DNA (a gene!) • We are testing to see which genes people have and if they have 0, 1, or 2 copies.

  7. What can it tell us? • If the gene contains the sequence CCCTTT, we can use a restriction enzyme that cuts at that sequence. • Person 1: ATCTATCTCCCTTTGGA • Person 2: AGTTGTAGTCCTTGATA Result = 2 fragments Result = 1 fragment

  8. Gel Electrophoresis Procedures • Get DNA sample and amplify it using PCR. • Mix DNA with restriction enzymes and allow DNA to be cut into fragments. • Pour buffer solution into electrophoresis machine (this will let the electricity flow) • Use a micropipettor to load DNA samples into small holes (called wells) in your gel (made of agarose)

  9. Micropipettor = used to pick up small, exact amounts of a sample You can set it to a certain amount and then it will pick up exactly that amount each time!

  10. Procedures continued . . . • 5) Place lid on machine and plug into power source. • 6) Set to the correct voltage and turn on. • 7) Let run until you see the sample has moved through the gel. • 8) Turn machine off and unplug it. Remove lid and take out gel. • 9) Stain gel with blue dye so we can see the fragments. • 10) Analyze gel.

  11. How does it work? • DNA molecules are slightly negative, so they are attracted to a positive charge. • The samples are placed at the negative end of the machine. • When the current is turned on, the DNA will move towards the other side of the machine because that side is positive. • Smaller fragments move faster than larger ones, so they are seen farther away from the wells. http://www.sumanasinc.com/webcontent/animations/content/gelelectrophoresis.html

  12. Analysis • People with lines (bands) in the same spots have the same genes. • Different positions tell us if the person has 2 copies of the dominant allele, two of the recessive allele, or one of each.

  13. Summary

  14. Go here to learn more! • http://learn.genetics.utah.edu/content/labs/gel/

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