S-STEM Scholar Leilani Boren's Blog

 Introduction:      Deinococcus geothermalis, the thermophilic bacterial species that has not one but two CRISPR systems according to genetic evidence, has been confirmed to have primers that are within range for acceptable efficiency levels so experimentation can begin. The original experiment included growing Deinococcus geothermalis and then exposing cultures to three different temperatures for a period of time to evaluate a change in gene expression of Cas2 and Cas10. This would potentially indicate that a change in environment can upregulate or downregulate CRISPR genes, which can be useful in understanding how these systems operate in life. It had been previously seen during primer efficiency validation runs that the baseline gene expression of D. geothermalis grown at a constant 45 degrees Celsius has a Cas2 and Cas10 transcription level that is similar to genes considered to be crucial for survival like our reference genes, Gap3 and SecA. This week, D. geothe...

 Introduction:     Last week there was a failed attempt at calculating the primer efficiencies of the Cas2 and Cas10 genes from D. geothermalis needed for the impending experiment to determine whether or not there is any change in gene expression at different growth temperatures over a 72 hour period. This week another attempt was made and failed at Cas2 and Cas10 due to over loaded cDNA from high yield RNA extractions, and finally the third attempt was successful in attaining favorable primer efficiencies for both Cas2 and Cas10 that indicates the experiment can move forward. As well as that, confirming the transformation of D. geothermalis with Prad1 was done with Prad1 AMP primers that would hopefully amplify a fragment 861 bp in length that could be visualized on a gel. The Prad1 primers were also verified to make sure they were working properly, and D. geothermalis tested in two ways.  Methods:     RNA was extracted from a culture of D. geothermalis gr...