Talk:CH391L/S13/GeneticMarkers

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***'''[[User:Aurko Dasgupta|Aurko Dasgupta]] 18:41, 7 March 2013 (EST)''': Are the products from Amp breakdown toxic to humans? Poisoning bacteria with something that turns into stronger poison doesn't sound all that bad otherwise.
***'''[[User:Aurko Dasgupta|Aurko Dasgupta]] 18:41, 7 March 2013 (EST)''': Are the products from Amp breakdown toxic to humans? Poisoning bacteria with something that turns into stronger poison doesn't sound all that bad otherwise.
**'''[[User:Catherine I. Mortensen|Catherine I. Mortensen]] 11:39, 7 March 2013 (EST)''':I've heard of some other antibiotics such as tetracycline, streptomycin, and chloramphenicol. When are these particularly useful?
**'''[[User:Catherine I. Mortensen|Catherine I. Mortensen]] 11:39, 7 March 2013 (EST)''':I've heard of some other antibiotics such as tetracycline, streptomycin, and chloramphenicol. When are these particularly useful?
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***'''[[User:Aurko Dasgupta|Aurko Dasgupta]] 18:38, 7 March 2013 (EST)''': The wikipedia page for [http://en.wikipedia.org/wiki/Streptomycin#Mechanism_of_action Strep] says that at low concentrations, it only inhibits growth instead of outright killing the cells. It cites a Biochemistry textbook for this. For Tet, the fact that it's counterselectable, as Larry mentioned, is extremely useful. According to [http://pharmaxchange.info/press/2011/05/mechanism-of-action-of-tetracyclines/ this], tetracycline binding to the 30S ribosomal subunit is reversible, which sounds useful, althoughI don't know how you'd actually reverse it.
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***'''[[User:Aurko Dasgupta|Aurko Dasgupta]] 18:38, 7 March 2013 (EST)''': The wikipedia page for [http://en.wikipedia.org/wiki/Streptomycin#Mechanism_of_action Strep] says that at low concentrations, it only inhibits growth instead of outright killing the cells. It cites a Biochemistry textbook for this. For Tet, the fact that it's counterselectable, as Ben mentioned, is extremely useful. According to [http://pharmaxchange.info/press/2011/05/mechanism-of-action-of-tetracyclines/ this], tetracycline binding to the 30S ribosomal subunit is reversible, which sounds useful, althoughI don't know how you'd actually reverse it.
*'''[[User:Gabriel Wu|Gabriel Wu]] 17:33, 4 March 2013 (EST)''': For those who work outside of E. coli, there's a class of selection markers used more commonly in yeast known as auxotrophic markers. Maybe not this time, but this topic should be addressed on this page at some point.
*'''[[User:Gabriel Wu|Gabriel Wu]] 17:33, 4 March 2013 (EST)''': For those who work outside of E. coli, there's a class of selection markers used more commonly in yeast known as auxotrophic markers. Maybe not this time, but this topic should be addressed on this page at some point.
**'''[[User:Benjamin Gilman|Benjamin Gilman]] 17:39, 5 March 2013 (EST)''': The [http://openwetware.org/wiki/CH391L/S12/Selectablegeneticmarkers selectable genetic markers] page from last year's class gives a brief overview of different categories of markers, including auxotrophic markers.  Briefly, auxotrophs are organisms that are incapable of synthesizing a particular molecule required for growth, in this case because they've had a necessary gene knocked out.  The mutant auxotrophs grow when supplemented with the required molecule but require complementation with the deleted gene to survive without it .  Auxotrophic markers are more often associated with yeast than bacteria mostly because the range of antibiotic resistance markers in yeast is much smaller.  The GFAT-expressing gene, ''glmS'', I discussed in class is an example of an auxotrophic marker which works in ''E. coli'', and a related GFAT gene, ''gfa1'', works similarly as a marker in fission yeast (''S. pombe'').
**'''[[User:Benjamin Gilman|Benjamin Gilman]] 17:39, 5 March 2013 (EST)''': The [http://openwetware.org/wiki/CH391L/S12/Selectablegeneticmarkers selectable genetic markers] page from last year's class gives a brief overview of different categories of markers, including auxotrophic markers.  Briefly, auxotrophs are organisms that are incapable of synthesizing a particular molecule required for growth, in this case because they've had a necessary gene knocked out.  The mutant auxotrophs grow when supplemented with the required molecule but require complementation with the deleted gene to survive without it .  Auxotrophic markers are more often associated with yeast than bacteria mostly because the range of antibiotic resistance markers in yeast is much smaller.  The GFAT-expressing gene, ''glmS'', I discussed in class is an example of an auxotrophic marker which works in ''E. coli'', and a related GFAT gene, ''gfa1'', works similarly as a marker in fission yeast (''S. pombe'').

Revision as of 18:43, 7 March 2013

  • Gabriel Wu 17:30, 4 March 2013 (EST): On a practical note, ampicillin (amp), kanamycin (kan), and chloramphenicol (cam) are the most used antibiotic selection markers in synthetic biology. Amp in my experience is convenient, but somewhat "fragile" (relative to kan and cam). Convenient: typically "rescuing" your cells post-transformation is unnecessary when using amp. "Rescuing" is necessary for kan and cam. Cam and kan are useful for long time course experiments (longer than 24 hours) because amp generally breaks down (or is broken down) over time.
    • Benjamin Gilman 17:53, 5 March 2013 (EST): Gabe brings up a good point, amp is an easy antibiotic to use because it doesn't kill the cells quickly, so you don't have to wait for beta-lactamase to build up before you plate them on antibiotic. Ampicillin is somewhat unstable, so many labs use it's relative cerbenicillin, which survives longer and breaks down into less toxic products. Ampicillin is cheaper though, and works pretty well most of the time, so it remains the popular choice.
      • Aurko Dasgupta 18:41, 7 March 2013 (EST): Are the products from Amp breakdown toxic to humans? Poisoning bacteria with something that turns into stronger poison doesn't sound all that bad otherwise.
    • Catherine I. Mortensen 11:39, 7 March 2013 (EST):I've heard of some other antibiotics such as tetracycline, streptomycin, and chloramphenicol. When are these particularly useful?
      • Aurko Dasgupta 18:38, 7 March 2013 (EST): The wikipedia page for Strep says that at low concentrations, it only inhibits growth instead of outright killing the cells. It cites a Biochemistry textbook for this. For Tet, the fact that it's counterselectable, as Ben mentioned, is extremely useful. According to this, tetracycline binding to the 30S ribosomal subunit is reversible, which sounds useful, althoughI don't know how you'd actually reverse it.
  • Gabriel Wu 17:33, 4 March 2013 (EST): For those who work outside of E. coli, there's a class of selection markers used more commonly in yeast known as auxotrophic markers. Maybe not this time, but this topic should be addressed on this page at some point.
    • Benjamin Gilman 17:39, 5 March 2013 (EST): The selectable genetic markers page from last year's class gives a brief overview of different categories of markers, including auxotrophic markers. Briefly, auxotrophs are organisms that are incapable of synthesizing a particular molecule required for growth, in this case because they've had a necessary gene knocked out. The mutant auxotrophs grow when supplemented with the required molecule but require complementation with the deleted gene to survive without it . Auxotrophic markers are more often associated with yeast than bacteria mostly because the range of antibiotic resistance markers in yeast is much smaller. The GFAT-expressing gene, glmS, I discussed in class is an example of an auxotrophic marker which works in E. coli, and a related GFAT gene, gfa1, works similarly as a marker in fission yeast (S. pombe).
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