User:Emilienne

=Student Registration/Questionnaire= Please copy the source code from this page to your user page (click on the button above, and copy everything inside the box to your user page). Fill in the answers and print out a copy for next time. You do not need to keep the information on your user page once you've printed it out.

Last Name
Repak

First Name
Emilienne

Course/Minor
Course 20, minor in French, strong interest in neuroscience

Year of Graduation
2009

Telephone #
(267)566-1418

Email
emilie7 at mit dot edu

Have you taken
7.05/5.07 (Biochemistry) currently taking 7.06 (Cell Biology) no 7.02 (General Biology Lab) no 5.310 (General Chemistry Lab) no

Do you have any experience culturing cells (mammalian, yeast or microbial)? No

Do you have any experience in molecular biology (electrophoresis, PCR, etc)? No

Please briefly describe any previous laboratory experience
UROP in Fee Lab in McGovern Institute, BCS. February 06- present. I do brain surgery on songbirds (lesions, tracers, and electrophysiology), and I perform perfusions, histology, immunochemistry, and microscopy.

I am currently exploring the role of the auditory cortex in song development, following up on some interesting previous results from a post-doc in my lab. An interesting side effect of the lesioning surgeries I do (in which the auditory input is removed from the auditory cortex) is that our birds become fearless. This might make another exciting follow-up experiment!

Anything else you would like us to know?
''Optional: As mentioned in lecture on 02.06.07, we would like you to consider how, as a biological engineer you might test the superstition "Spit on a bat before using it for the 1st time to make it lucky." ''

I would perform a double blind experiment in which both athletes testing bats and evaluators scoring performance of the bats do not know which bats have been spit upon. One could then statistically analyze the data from bats that had and had not been spit upon to see if bats with spit on them were any luckier. If, as I suspect, they are not, then I have proven that I do not need to go into molecular analysis of the spit to try to find the elements which contribute to the luck.

=M13 Genome Engineering=

M13 Design Constraints
Would you expect the phage to tolerate p8 modifications that: 1. make the protein neutral rather than negatively charged at the C-terminus?

It seems that the C-terminus may need to be charged in order to hold together the P8 proteins that make the phage coat. The charge may also be necessary for intermolecular interaction required to secrete the phage. It seems that a neutral protein will be less likely to have intermolecular interactions than a negatively charged protein, so I think that the phage would struggle to tolerate this modification. (I tried learning about the C-terminus of P8 from the abstract at the following site: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=12217696&dopt=Abstract )

2. encode all the Leucines with the CTA codon instead of the CTG codon?

The codon sequence is not important as long as the appropriate amino acid is encoded, so the phage could tolerate this modification.

3. double the size of the protein?

It would be useful to know how the p8 knows how to regulate its production. Since the amount of p8 produced decreases when the genome size is decreased, this leads me to suspect that somehow the amount of p8 is being regulated such that the phage coat will always fit snugly over the phage’s DNA. Therefore, if the size of p8 is doubled, I suspect that only half the amount of protein will be used and the phage will tolerate this change just fine.

Would you expect the phage to tolerate these same modifications to p3?

1. C-terminal domain is required for release from the host cell following phage assembly, and contributes to structural stability of phage particle (See abstract http://chem.ps.uci.edu/~gweiss/WeissG-03JMB.pdf ). If we change the charge, it will alter intermolecular interactions, and it may destroy the functionality of p3, thus preventing the release of the phage and/or destabilizing the phage.

2. The codon sequence is not important as long as the appropriate amino acid is encoded, so the phage could tolerate this modification.

3. If the C-terminal is doubled in size, I think that it will still function normally, though it’s possible that it will not be able to pass through the P1-P4-P11 complex through which mature phage are secreted.

Would you expect the phage to tolerate transcriptional terminators that are 1. 2X stronger 2. 100X stronger 3. 2X weaker 4. 100X weaker ?

Assuming that we want the transcription to be stopped at a transcriptional terminator, it seems that it wouldn’t be a problem if the transcriptional terminator was stronger, unless an excessively strong transcriptional terminator caused the end part of the protein to be cut off. If a weak transcriptional terminator causes a protein to have a long tail that could potentially cause problems, should the tail undergo undesired intermolecular interactions of simply take up too much space in the phage. It may also be possible that a weak transcriptional terminator would not successfully stop translation of the protein, which would be a major problem.

The Family Tree: M13's closest evolutionary relatives
M13 is a bacteriophage in the family Inoviridae, known for being simple, circular single-stranded-DNA filamentous phage with very similar genetic design. Two of M13's closest relatives are bacteriophage fd and fl. Both of these phage differ from M13 slightly in their coats. M13 has an amide group of asparagine in position 12 whereas fd and fl have a carboxyl group of aspartate. (See http://www.wsu.edu/~hipps/pdf_files/phagep1.pdf ). "As a result of this difference, M13 has one negative charge less per subunit than fd, corresponding to a 25-30% reduction in surface charge density" (Tang et al., “Metal Ion-Induced Lateral Aggregation of Filamentous Viruses fd and M13”. http://www.biophysj.org/cgi/content/full/83/1/566 )

For further information on bacteriophage and a useful comparison of 12 different catergories of phage, see http://www.tulane.edu/~dmsander/WWW/224/Phages.html.