User:Pakpoom Subsoontorn/Notebook/Genetically Encoded Memory/2008/10/06: Difference between revisions

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* In state A, the cell has two pieces of DNA, the chromosomal DNA and  
* In state A, the cell has two pieces of DNA, the chromosomal DNA and  
the free circular plasmid. The chromosomal DNA has two different bacterial integration  
the free circular plasmid. The chromosomal DNA has two different bacterial  
sites for integrase, attB-1 and attB-2. The plasmid also has
integration sites for integrase, attB-1 and attB-2. The plasmid also has
two different phage-integration sites for integrase, attP-1 and attP-2.
two different phage-integration sites for integrase, attP-1 and attP-2.


Line 20: Line 20:
state-C  
state-C  


    *  These are some ideas about reversible genetic knockout
    * Can we have a system of two pieces of DNA that can fuse and re-split? The fusion is triggered by signal-1, splitting is triggered by signal-2. Well, this will be similar to the way phages enter and exit their lysogenic cycle. Now, the challenges are the following. First, unlike natural phage, the split of DNA must not lead to the lytic cycle. We want to host to survive. Second, the host has to maintain the two separated pieces of DNA. Third, the fusion and splitting efficiency must be high.
    * Can we modify integrase system to function like transposon (but higher specificity/ efficiency)? The piece of jumping DNA of attP for two different integrase systems. Signal-1 makes it insert to attB-1; signal-1' cuts it out; signal-2 makes it insert at attB; signal-2' cut it out again.
    * Readmore about transgenic/ knockout organism. How to target a specific gene in a genome? Read Zebrafish knockout paper


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Revision as of 20:24, 6 October 2008

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mini-Project: Reversible Genetic Knockout

  • The genetic systems has two states: A, B, C. The state can be changed

upon the expression of two different intergrase/excisionase systems.

  • In state A, the cell has two pieces of DNA, the chromosomal DNA and

the free circular plasmid. The chromosomal DNA has two different bacterial integration sites for integrase, attB-1 and attB-2. The plasmid also has two different phage-integration sites for integrase, attP-1 and attP-2.

  • The expression of the integrase-1 will insert attP-1 into attB-1, changing

the system from state-A to state-B. On the other hand, the expression of integrase-2 will insert attP-2 into attB-2, changing the system from state-A to state-C


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Management Questions/requests

  • Should we push the review article on Genetically Encoded Memory to general

audiences? or just leave it as a note among ourselves?

  • As we discussed earlier, there are 3-4 levels of the project: The application level (like engineering a yeast to report its age, or an in vivo directed evolution system), the grand-challenge level (like making 8-bit counter) and the specific experiment level. How should we manage our time?
  • Relating to the question above, should I start a deep-research on a particular system (say, lambda integrase)? or should I explore broader ranges of tools (other enzymes, recombination system etc.) for now?

Science Questions/ To-do list

  • These are some ideas about reversible genetic knockout
  • Can we have a system of two pieces of DNA that can fuse and re-split? The fusion is triggered by signal-1, splitting is triggered by signal-2. Well, this will be similar to the way phages enter and exit their lysogenic cycle. Now, the challenges are the following. First, unlike natural phage, the split of DNA must not lead to the lytic cycle. We want to host to survive. Second, the host has to maintain the two separated pieces of DNA. Third, the fusion and splitting efficiency must be high.
  • Can we modify integrase system to function like transposon (but higher specificity/ efficiency)? The piece of jumping DNA of attP for two different integrase systems. Signal-1 makes it insert to attB-1; signal-1' cuts it out; signal-2 makes it insert at attB; signal-2' cut it out again.
  • Readmore about transgenic/ knockout organism. How to target a specific gene in a genome? Read Zebrafish knockout paper

References


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