Science Buddies: "Ask an Expert"

To Whom It May Concern:

I am a student doing a project on detmining the phylogenic relationships of different vegetables by comparing their genomes using gel electrophoresis. I have the procedure for the electrophoresis, but I am unsure of the procedure for extracting the DNA from the cells of the specimens. I also would like to know whether or not i need to use PCR to clone the DNA for the electrophoresis, or if I can get enough DNA just from the vegetables themselves.

Thank you for your aide. It is greatly appreciated.

Sincerely,

Kelsey Daly

Kelsey,
you won't be able to diretly compare genomes by electrophoresis of extracted DNA- there are several ways to make comparisons by gel analysis: two options are 1) cut the genomic DNA with a series of restriction enzymyes and analyze the fragments by electrophoresis (agarose gels); 2) PCR specific regions of the genome and analyze amplified fragment size

one other thing to keep in mind is that agarose gels are usually visualized by ethidium bromide and UV light- both are dangerous. ethidium bromide is a carcinogen and needs to be disposed of correctly.

As for making DNA extracts, I assume you have access to the tools required. There are protocols are available on the web and from many lab web pages. A few places to start:

http://www.ucs.louisiana.edu/~mrv8332/H ... plants.doc

http://www.sidwell.edu/us/science/vlb5/ ... n_lab.html

http://pubs.nrc-cnrc.gc.ca/ispmb/ispmb17/17053-1.pdf


You might also consider a bioinformatics approach- many plant sequences are available in public databases, and the degree of homology between either protein or DNA sequence is realtively easy to determine using freely-available tools.

Kelsey:

The Sciencebuddies home page has a section called Cutting Edge Projects. There is a wealth of genome data available from public sources.

Experts,

I do indeed have the equipment to perform the DNA extraction. Thank yuo for this information. I intend to use restriction enzymes and measure the differences in the location of the DNA. Do you think that I will need to perform PCR in order to get enough DNA to perform the electrophoresis?

Thank you,

Kelsey Daly

Yep, you can extract enough to perform digests without amplifying, although you'll probably need to use a range of enzymes to get a good picture of species diversity- start with the 8 cutters and move down. Also, you'll probably need to use different extraction methods for high-yield recovery of different plants: high-starch veggies tend to require different protocols than the others.

best of luck,
htb