Talk:CH391L/S13/Riboswitches

• Catherine I. Mortensen 13:45, 25 February 2013 (EST): How practical is DNT detecting E. Coli in airports and other public areas considering the time it takes for E. Coli to start producing fluorescent protein?
  • Benjamin Gilman 18:15, 28 February 2013 (EST): For that application it's not useful at all, but in a previous paper they used a fluorescently tagged version of the same aptamer to detect TNT in vitro in a purpose-built instrument Biosensor-based on-site explosives detection using aptamers as recognition elements. The aptamer was generated to bind TNT, but DNT was used in the later paper because it's easier to obtain and still binds well. The E.coli system may be more sensitive for detecting small amounts of DNT or TNT in things like groundwater, where they're using it to detect contamination and the speed of the test doesn't matter as much.

• Kevin Baldridge 17:08, 25 February 2013 (EST):Are there any known examples of a riboswitch with an IRES (Internal Ribosome Entry Site, an alternate way to initiate translation that doesn't use the Shine Dalgarno sequence) expression platform? If not, that might be something we could look into for iGEM, directed evolution or SELEX to design such a system.
  • Catherine I. Mortensen 08:59, 26 February 2013 (EST):What would be the advantage of IRES as opposed to a Shine Dalgarno?
    • Benjamin Gilman 18:36, 28 February 2013 (EST): They're used for different application. Shine-Dalgarno sequences function in translation initiation in bacteria and archaea, whereas IRES's are generally used to initiate translation in eukaryotes, bypassing much of the cell's translation regulation machinery.
  • Catherine I. Mortensen 19:39, 28 February 2013 (EST):I know there is a lot of research being done with aptamers that bind to cancer cells so that the immune system has a better way of recognizing cancerous cells
  • Benjamin Gilman 18:36, 28 February 2013 (EST): A researcher named Atsushi Ogawa has published two papers about using a riboswitch to control an IRES as either an ON switch or OFF Switch in response to theophylline.
    • Catherine I. Mortensen 19:48, 28 February 2013 (EST): UCLA seems to be working on an aptamer that inhibits a type of HIV virus from infecting the cell [1]

• Gabriel Wu 17:17, 25 February 2013 (EST): Connecting with Max's topic, directed evolution strategies are very relevant in developing riboswitches, especially for aptamer design. Might be nice to discuss the connection between the two topics.
  • Max E. Rubinson 22:21, 28 February 2013 (EST): An old professor of mine at Emory, Justin Gallivan, uses high-throughput selection strategies to identify synthetic riboswitches constructed from RNA aptamers.
    • Max E. Rubinson 22:26, 28 February 2013 (EST): Here is a link to a book chapter discussing the selection methods they use.
• Kevin Baldridge 17:18, 25 February 2013 (EST):Here is a nice example of taking advantage of riboswitch concepts for synthetic biology Isaacs 2004, our group is working on developing a probe for in vivo analysis of RNA tertiary structure
  • Gabriel Wu 21:54, 28 February 2013 (EST): A year later, Bayer and Smolke showed that you could use aptamers to regulate a riboswitch in yeast.
• Kevin Baldridge 17:26, 25 February 2013 (EST):it looks like your biblio section might have a few of the entries merged together at the end?
  • Catherine I. Mortensen 08:59, 26 February 2013 (EST): Thank you! Fixed.
• Catherine I. Mortensen 19:24, 28 February 2013 (EST): Aerobic bacteria are one of the causes of spontaneous combustion of hay stack, manure, and compost piles. What if an eGFP riboswitch was added to these bacteria so that the more they oxidized their surroundings, the more fluorescent protein they would create. Areas of big manure or compost piles would light up in areas vulnerable to catching on fire.This way farmers could better control hay fires and such by noticing the amount of fluorescent protein present. Is this plausible....?
• Alvaro E. Rodriguez M. 22:13, 28 February 2013 (EST): You could also briefly mention what the concept of attenuation means...as part of the historical background and google TPP and IGEM as this relevant connection.
  • Catherine I. Mortensen 12:04, 4 March 2013 (EST): TPP works at the post transcriptional level as a transcription factor. Can this be considered a ligand in the context of a riboswitch?
• Thomas Wall 23:17, 28 February 2013 (EST): E. coli don't have native cobalamin synthesis, people have incorporated the pathway from salmonella into e. coli but it is a completely anaerobic process. Does anyone know if this an enzymatic requirement or a genetically controlled event? I was thinking that we could use the cobalamin riboswitch to monitor an aerobically or anaerobically functioning synthesis pathway. Also, I believe this would be an industrially relevant process as B12 is needed as a cofactor for many processes.
• Thomas Wall 23:30, 28 February 2013 (EST): I'm dumb when in comes to riboswitches, is there any reliable computer method for predicting them in the genome? Also it seems like something generalizable to turn an aptamer into a riboswitch needs to be figured out, or maybe it has?
• Yunle Huang 00:19, 1 March 2013 (EST): Could someone clarify why the polymerase falls off when the hairpin forms? Does the hairpin formation reveal the polyuracil region?
  • • Catherine I. Mortensen 13:46, 1 March 2013 (EST): The presence of a polyuracil region is not dependent on whether or not the hairpin forms. A polymerase will transcribe through a polyuracil region but the speed at which it does this decreases because something about the chemistry of uracil (possibly due to the fact that uracil can only form 2 hydrogen bonds instead of 3 like guanine and cytosine can) slightly destabilizes the polymerase. When the hairpin forms, the polymerase becomes too unstable to hold onto the polyuracil region. It's as if the polymerase runs over a bump and loses control I'd say.
• Siddharth Das 00:58, 1 March 2013 (EST): Can riboswitches be synthesized chemically? Correct me if I'm wrong, but I find it interesting that the rational design of riboswitches could provide a means of mediating gene expression in other organisms. What does this "promiscuity" translate to in terms of an evolutionary standpoint?
  • Catherine I. Mortensen 13:58, 1 March 2013 (EST):I know there is a lot of research being done with eukaryotes and riboswitches but most of the papers I came across were focused on designing a ligands but I didn't come across any completely synthetically made riboswitches in eukaryotes.

• Marco Howard What is the time scale for riboswitche to function. For example, if you use them them as sensors how long does it take to get a positive or negative signal. Also, if you are using them to express a gene, how long does it take for a protein to folded and functional?]
  • Catherine I. Mortensen 13:38, 1 March 2013 (EST): A lot of graphs are measured in hours so I assume hours...