User:Chaoj
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20.109 Spring 2007
Last Name
Chao
First Name
Jennifer
Preferred name
Jen
Course/Minor
Course 20, minor in Management
Year of Graduation
2009
Telephone #
610-952-3098
chaoj at mit dot edu
Day 1: Start up Genome Engineering
| protein | function | re-engineering ideas |
|---|---|---|
| I | assembly | p1 and p11 interact with p4 so within the bacterial inner membrane, modify p1 to make secretion of the phages more effective. (speed/size of channel) |
| II | replication of DNA + strand | modify p2 so it nicks in such a way to make p5 respond quicker, send out a signal to p5 so it can protect the + stranded DNA |
| III | phage tail protein (5 copies) | add a tag to monitor phage escaping from host and the new phages budding from the bacterial surface |
| IV | assembly | modify p4 so assembly speed is increased in the outer membrane and modify it so it has a better interaction with p1. This will cause the channels where the phages are secreted to be more effective |
| V | binds ssDNA | modify p5 so it can better protect the + stranded DNA which will lead to more rapid replication and amplification; less competition with the double stranded DNA |
| VI | phage tail protein (5 copies) | similar to p3, p6 is the accessory protein to p3. Modify it so it has better and more effective interaction with 3, no interference |
| VII | phage head protein (5 copies) | companion protein to p9, make sure or modify p7 so it can better the interaction with p9, better binding affinity. |
| VIII | phage coat protein (2700 copies) | Modify so the shape changes by altering the coat formulation, make the phage more streamlined, sleek so increases proliferation speed. |
| IX | phage head protein (5 copies) | Modify resulting in faster secretion of DNA |
| X | DNA replication | linked to p2, modify so p10 is not so dependent on 2 and the DNA can still replicate and modify so there is better accumulation of + stranded DNA |
| XI | assembly | modify similar to p1 to change channel properties to make secretion of phages more effective |
Day 4: Genome Refactoring
I refactored the phage genome to remove overlaps and added in restriction enzyme sites, which EcoRV and HindIII cut, between each part, more specifically Gene 8 and the Gene 3 promoter. I changed a few bases, and consequently some codons, but retained the same amino acid sequence while weakening the rbs or promoter. Considerations: 1)Unstuffed by splicing all genes from smaller parts into composite parts; removed overlaps. 2) eliminated promoter of gene 3, by changing a few bases that would not change amino acid sequences but would decrease the strength of the rbs or promoter. 3) add restriction sites between each part BBa_M30058.
New Parts: restriction enzyme sites
| restriction enzyme | code |
|---|---|
| EcoRV | ATC |
| HindIII | A^AGCT_T |
