S-STEM Scholar Leilani Boren's Blog

 Introduction: In previous weeks Illumina sequencing has been well underway, with multiple species and some isolates have newly developed de novo sequences that has provided a plethora of information and insight. This week was a continuation and preparation for the following week in which more isolates and species will be sequenced, including a replication of D. caeni due to incomplete data acquired the first run on Illumina. The two isolates chosen are considered to be 16s outliers in comparison to other isolates, and suspected of being the most genetically variant from type strain of D. aquaticus, PB314. The other two species, D. pimensis and D. grandis were chosen due to only having limited sequencing data available. D. pimensis has only scaffolding information and D. grandis has contig files is listed as having high levels of contamination that can and should be improved upon for clarity. This week included ensuring the quality of the stored DNA samples, cleaning up a few of th...

Introduction: Deinococcus sonorensis is a species of the genus that has been intensively studied in this lab previously due to its abnormal and unique characteristics when it is grown on TGY media. A poster presentation to develop an optimal DNA extraction method even when it forms its plaque structure was completed about a year ago, and now it is time to revisit the advancements that have been made in the past six months. Fortunately Illumina sequencing became available and the mysteries of D. sonorensis has started to unravel, thanks to being able to bioinformatically process this species and the addition of biochemical testing that has shed light on some new ideas. Not only are we now able to grow D. sonorensis on media and prevent plaque formations, we can extract its DNA better than ever before, and have findings that allow us to infer what proteins or systems may be responsible for its odd behaviors that make this species so interesting in the genus.  Methods: D. sonorensis w...

  Introduction: The 7 kb plasmid of Deinococcus aquaticus has been a long term goal for the lab to not only isolate effectively, but to genetically engineer in a way that it may potentially be useful for other Deinococcus applications in the future. The roadblocks with this plasmid began in just extraction methods from D. aquaticus, with those efforts not resulting in any significant clarity in how to most efficiently isolate this plasmid from living cells in high enough quantities to work with. Thus the idea to amplify the plasmid in a long amp procedure came to be, in which primers were made by Jonathan to not only produce considerable amounts of the plasmid but equip it with long overhangs of nucleotides that would be compatible for the Gibson assembly. So far there has been considerable difficulty validating this procedure in order to put chloramphenicol resistance (Cmr) onto the 7 kb so that this new plasmid could be tested in an array of species in the genus. This week, progr...