1. iGEM Meeting #4 07/16/08 Presentation by Robert Ovadia
Lactose Intolerance
iGEM Meeting #4
07/16/08
Presentation by Robert Ovadia
I chose this background because it looks like it had two plasmids. It relates to my project because to solve lactose intolerance, we need two plasmids. The first being *click*

2. iGEM Meeting #4 07/16/08 Presentation by Robert Ovadia
System Design
The idea behind using RBS’s is we want to use the weakest RBS for lacY so we can pump the strongest promoter. We want to express LacZ as much in order to do this.
*ADD ORI’s*
iGEM Meeting #4 07/16/08 Presentation by Robert Ovadia

3. iGEM Meeting #4 07/16/08 Presentation by Robert Ovadia
System Design
By next week, you guys should be telling me how the plasmids look like. I will test you!
*ORI’s*
iGEM Meeting #4 07/16/08 Presentation by Robert Ovadia

4. Last Week’s Results What we had
We have sequenced:
We have transformed:
LacY 34
LacZ
Into pSB1A2
LacY
Purified ready to cut:
From PCR
Lysis Cassette
We will send another one in for sequencing to determine what really happened.
There was a single base mutation. It would have been nice if it didn’t change the amino acid, but unfortunately it did. We are hesitant to use this construct because 1 mutation could alter the folding of the protein and the function.
So all we have now is LacY in pSB1A2
*MAKE SIMPLER*
PCR Purified 31
LacY 31 33 33
LacY
iGEM Meeting #4 07/16/08 Presentation by Robert Ovadia

5. Last Week’s Results Sequencing: The Work Chart
Did it work?  31
LacY  33
LacY  34
LacZ
We will send another one in for sequencing to determine what really happened.
There was a single base mutation. It would have been nice if it didn’t change the amino acid, but unfortunately it did. We are hesitant to use this construct because 1 mutation could alter the folding of the protein and the function.
So all we have now is LacY in pSB1A2
*MAKE SIMPLER* 
Lysis Cassette
iGEM Meeting #4 07/16/08 Presentation by Robert Ovadia

6. Future Goals What’s next?34
LacZ
In J23100 Str. Po.
Lysis Cassette
In pSB1A2
In pSB2K3 31
LacY
We have our three constructs that we currently have. Now what do we want to do this them for next week?
LacY into 31/33, the weaker RBS’s 0.01 and 0.07 (7 fold increase). From 33 to 32, 4 fold increase.
LacY + LacZ into TT
Primer design for mutagensis
Lysis cassette into pSB1AK3. Photo of lysis cassette.
*ADD MORE*
In pSB1AK3 33
LacY
In pSB1AK3
iGEM Meeting #4 07/16/08 Presentation by Robert Ovadia

7. iGEM Meeting #4 07/16/08 Presentation by Robert Ovadia
Current Constructs
We have sequenced:
Preparing to digest:
A whole lot of stuff…
Three promoters
Lac. inducible promoters (A81, B4)
RBS, GFP ,TT
Our current constructs
B0015 (1AK3 + 2K3)
LacY 31
LacY 33
LacY
*UPDATE* 34
LacZ
Lysis Cassette
iGEM Meeting #4 07/16/08 Presentation by Robert Ovadia