COMPARISON OF RIBOSWITCH REPORTER SYSTEMS FOR LIVE CELL IMAGING OF CYCLIC-DI-GMP DYNAMICS IN BACILLUS SUBTILIS POPULATIONS
| dc.contributor.advisor | Spirito, Catherine | |
| dc.contributor.author | Pierre, Yasmine | |
| dc.date.accessioned | 2020-04-27T02:54:23Z | |
| dc.date.available | 2020-04-27T02:54:23Z | |
| dc.date.issued | 2019 | |
| dc.description.abstract | Fluorescent riboswitch reporters can be used in vivo to monitor metabolite dynamics. Previous work used a fluorescent yfp reporter based on a cyclic di-GMP responsive riboswitch from Bacillus licheniformis to monitor cyclic di-GMP levels in individual Bacillus subtilis cells. The previous study found that cell fates in Bacillus subtilis are not uniform in the presence of varying cyclic di-GMP levels. It is important to further develop tools that enable single-cell imaging in Gram-positive bacteria. Fluorogenic aptamers are single-stranded RNA molecules that have been evolved via in vitro selection to bind strongly and specifically to fluorophore molecules and emit a fluorescent signal. These fluorogenic aptamers can be used instead of fluorescent proteins in riboswitch reporter systems to provide a more dynamic read-out of metabolite dynamics in cells. However, relatively little work has been done to evaluate the use of these fluorogenic aptamers as reporter systems in Gram-positive bacteria. The objective of this project is to evaluate the use of four different fluorogenic aptamers (Mango-III, Broccoli, dimeric Broccoli, and SpinachII) instead of yfp in a cyclic di-GMP responsive riboswitch reporter system in Bacillus subtilis. All plasmids containing the riboswitch reporters were constructed and successfully transformed into E. coli cells. Subsequently, the cyclic di-GMP responsive riboswitch reporter systems were successfully transformed into B. subtilis WT PY79 and a 𝝙pdeH mutant. Future work involves evaluating their performance in vivo in B. subtilis via laser confocal and fluorescence microscopy. | en_US |
| dc.description.sponsorship | FIRE LSAMP NSF | en_US |
| dc.identifier | https://doi.org/10.13016/1zxw-eh7g | |
| dc.identifier.uri | http://hdl.handle.net/1903/25919 | |
| dc.relation.isAvailableAt | Maryland Center for Undergraduate Research | |
| dc.relation.isAvailableAt | Digital Repository at the University of Maryland | |
| dc.relation.isAvailableAt | University of Maryland (College Park, Md) | |
| dc.subject | FIRE | en_US |
| dc.subject | Pierre | en_US |
| dc.subject | LSAMP | en_US |
| dc.subject | Fluorogenic Aptamer | en_US |
| dc.subject | Single Cell Imaging | en_US |
| dc.title | COMPARISON OF RIBOSWITCH REPORTER SYSTEMS FOR LIVE CELL IMAGING OF CYCLIC-DI-GMP DYNAMICS IN BACILLUS SUBTILIS POPULATIONS | en_US |
| dc.type | Presentation | en_US |