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

From OpenWetWare
Jump to navigationJump to search
Line 6: Line 6:
| colspan="2"|
| colspan="2"|
<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->
<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->
==Notes and Thoughts==
==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 site for integrase, attP-1 and attP-2.
 
 
*'''Bold text'''
*'''Bold text'''
* Φc31 (and other serine recombinase) seems to be good choices for modify DNA code in term of efficiency and simplicity. They need only ~30 sth recognition sites on the inserting and the target DNA; no host recombination factor seems to be required; it has been shown to function in vitro and in non-natural host ...like animal and plant cell.  
* Φc31 (and other serine recombinase) seems to be good choices for modify DNA code in term of efficiency and simplicity. They need only ~30 sth recognition sites on the inserting and the target DNA; no host recombination factor seems to be required; it has been shown to function in vitro and in non-natural host ...like animal and plant cell.  

Revision as of 20:12, 6 October 2008

Project name <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page
<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>

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 site for integrase, attP-1 and attP-2.


  • Bold text
  • Φc31 (and other serine recombinase) seems to be good choices for modify DNA code in term of efficiency and simplicity. They need only ~30 sth recognition sites on the inserting and the target DNA; no host recombination factor seems to be required; it has been shown to function in vitro and in non-natural host ...like animal and plant cell.
  • One drawback is that their structures are poorly known and there is no known excision system yet.
   *  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

[edit]

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


Retrieved from "https://openwetware.org/mediawiki/index.php?title=User:Pakpoom_Subsoontorn/Notebook/Genetically_Encoded_Memory/2008/10/06&oldid=249295"