S-STEM Scholar Leilani Boren's Blog

 Introduction:     After weeks of trying to adjust the protocol to optimize the DNA extraction process, it was decided this week that the best option would be to attempt to transfer the original samples from last summer that are currently in TE buffer, as discussed previously is unsuitable for sequencing since it contains trace amounts of EDTA, to water. The fear in doing this is that some samples will fall out, predominantly by not having enough DNA input to the ZYMO DNA cleanup and concentrator kit which has been used in the past. It seems as though if some specie samples do fall out the best course of action will be to go back to bead beating lysis methods with the DNeasy kit by QIAGEN, but we would like to avoid this as much as possible to prevent even further issues with sample extraction and time. Unfortunately time is a critical factor because the sequencing equipment is only useful for a certain number of days before it is ineffective for sequencing, so the fewer ...

 Introduction: Many previous weeks of work has been dedicated to developing DNA extraction protocols for third generation sequencing in order to produce data that will give insight to Deinoccocus species that have been neglected by the scientific community thus far. Last summer the method of choice after lengthy testing and the urgency to use a protocol for high molecular weight (QIAGEN Genomic tip protocol) was decided to be sonication for the lysing of cells because according to previous gels these samples produced high molecular weight and visible bands greater than 45kb, the highest seen in this lab to date. This week, we attempted to sequence two different species. The first. Deinococcus sonorensis was originally isolated by sonication and the lysate directly cleaned with the ZYMO DNA cleanup and concentration kit, which did not consistently work for other species after these runs. The second specie, Deinococcus pimensis was isolated as described from last week, with sonicat...