Koeng

While cross-referencing data from several different vectors, I remember Lambda’s mechanism for replication. It hides until the host’s chromosome is disrupted by U.V. light. While I was thinking of this, and how big it’s chromosome was, I realized that it could be possible to cut the gene for repressing the lambda’s genome into a plasmid (pUC18 or 19 for high plasmid yield), along with it’s bactericide. Maybe a few modifications here and there. This could be a high yield plasmid, and unlike lets say a phix174 vector, no need for taking out the viral proteins. Once they where in a vector, when you wanted to get the plasmids back no need for the complicated plasmid removal, just use U.V. light for a few hours and once they are lysed, extract the plasmids by say, centrifugation! I am no expert at how to extract plasmids, but does anyone think this might be a good idea? (p.s. anyone know any archaeal plasmids?)

Good-

No more buying plasmid extraction kits!
Possible to keep plasmids in bacteria and use other methods with experimentation
Make up your own idea!

Bad-

Why not just use lambda?
Why not just use another viral vector with the proteins taken out?
This would be hard to make!

Any input?

-Koeng

-(Edited Portion) I could make a map with either SnapGene viewer or SerialCloner if anyone wants

Hello! I am bored this summer, so I decided to make a hypothetical synthetic plasmid sequence. I wanted this plasmid to be as SMALL as possible. It took quite a while but I completed a possible plasmid. This plasmid is based off the pUC19 vector. I took out the lacZ gene since I saw it as unnecessary, even though it might turn the bacteria blue. Another reason I took out the lacZ gene is because I want just ampicillin in the agar plates for transformation because it is cheaper . I also saw that there was quite a bit of space in-between the genes, and since a lot of this is useless, I took it out. I still wanted some spots for restrictive enzyme digestion, so I put those in-between the origin of replication and the "bla" gene (for ampicillin resistance). I wanted those enzymes to be easy to get, so I choose Hindlll, BamHI, and EcoR1, which are in the DNA in about the middle, the sequence fragment is gaattcggatccaagctt. If there are any nice, short Multiple Cloning sites from any other plasmids that I could add in would be greatly appreciated.  I am no expert on protomers, but I believe the ones in the original sequence would work so I didn't change their spots. Any input would be great!

The ampicillin resistance gene (bla) is 641 - 1501and the origin of replication is 1 - 553. The "bla" protomer is 572 - 576 (TTCAAA) and the other "bla" protomer is 592 - 597 (GAGACA). I set it up on ApE (a plasmid Editor) but I can't upload that type of file, nor can I upload a map . But I did put the sequence in a nice text edit file. Thank you in advance!!!

-Koeng