Hi Jessica,
The mitochondrial DNA results are on
First, you should have all the students register online for the “sequence server” on the main page. This will allow the students to keep the data on their workspace without having to retrieve it every time they log in.
After registering, go to bioservers.org and sign into "sequence server". Go to "manage groups" and click on "classes" as a sequence source. Your samples are under my name and the3/5/15date. The tracking number is10-289862049.
If you click on the “view” button to the right of the row, the students should be able to see the chromatograms without downloading any software. Here, students can select their data from a drop-down list (the number before the "lcmtf" is the number assigned to each sample. All of the samples kept the same # as they were in class). When students look at the chromatograms, some students will notice that the quality of their sequences decreases after a homopolymeric region of DNA sequence. These are student #s4, 5, 8, 21, 22, 27, 29, and 30.These samples have a string of C's near the 190th nucleotide. You'll notice that after this direct repeat of C's that the sequence is of poor quality, due primarily to DNA polymerase slippage in the sequencing reaction. Everyone else in theclass has a slightly different sequence of DNA. You'll see most, if not all, of the other samples have a "T" within thisrepetitive C region. This minimizes DNA polymerase slippage and results in good quality sequence after this region.
To get the sequences on your workspace, click on the box to the left of the tracking number. Then click on the "OK" button on the bottom. Once you do this, all the sequences you selected are going to be on your workspace. Here students can then compare their sequences to each other.
Each student should calculate the % difference between him/her and another student. To do this, the students will first need to trim the sequences (trim the yellows at the beginning and end of the comparisons) and then use the following formula:
% difference = ______#mutations (yellow areas)______x 100
Total # bases compared (minus gray base comparisons)
Please note: A big chunk of yellow bases together in a high quality sequence comparison should be counted as one mutation (most likely due to a larger insertion or deletion). Gray areas indicate nucleotides that could not be determined in the sequencing reaction. They are designated as “N”s (“N” means “nucleotide” [it could be A, T, G or C!]). “N”s should not be counted as mutations or in the total # of bases compared.
The average difference between any two people in this mitochondrial DNA region is 2%.
When student #4, 5, 8, 21, 22, 27, 29, or 30compares his/her DNA to anyone else, he/shemay notice a lot of yellow bars after this string of C’s, even after trimming the comparisons. In this case the students should not count these nucleotides in the total # of bases calculation due the poor sequence quality. They can probably only count the bases that lead up to the string of C’s in their sequences. This is also true for anyone else in the class who compares their DNA to anyone with this homopolymeric region.
One student unfortunately did not get a good quality sequence at first. This is student #3. I asked Genewiz to do a repeat run of the sample and the repeat was of much better quality. Make sure that student #3 selects sample
Midwood_HS_030315-03-lcmtf_R at the bottom of the drop down list AND NOT Midwood_HS_030315-03-lcmtf.
It is a great idea to not only upload class data, but data we have on the server. On the “Manage Groups” page you can choose different sequence sources (e.g. modern human mtDNA or Neanderthal mtDNA) for comparisons. Make sure that when you go to your workspace to compare sequences that you check only the sequences you want to compare. Uncheck any data that you do not want to compare.
If you have any questions, check out the instructions that pop up after logging in. If that doesn’t answer your question, please feel free to ask me by email or phone.
Good luck and have fun!
Melissa :)