Progress 03/15/18 to 03/14/20
Outputs
Target Audience:Research scientists Changes/Problems:The student successfully validated the methodology for isolating DNA from mouse fecal pellet test samples, and we determined that processing of the more limited samples (approximately 1100 samples consisting of 1-2 fecal pellets per mouse) from the multigenerational study could proceed. However, this protocol proved unreliable on the large scale processing with fewer than 30% of samples passing quality control checks. Because these samples were randomized, it was also determined that an insufficient number of samples for experimental groups were available and we could not justify proceeding with the sequencing, as no meaningful, interpretable data could be obtained. Unfortunately, insufficient starting material was available to go back and start again. What opportunities for training and professional development has the project provided?During the fellowship training period, the graduate student participated in many professional development events and other training opportunities. For example, she has routinely attended the Graduate Research Training Series at USU; a lecture on rethinking science communication; workshops on resume's/cover letters and job negotiations, and other relevant webinars and colloquiums. She has also been active within the Society of Toxicology and American College of Toxicology organizations as a student leader. How have the results been disseminated to communities of interest?The student funded on this fellowship attended many conferences to present her doctoral research including: Environmental Mutagenesis Society (2018), the American Society for Nutrition annual meeting (2018, 2019), the American College of Toxicology (2018) and the Society of Toxicology annual meeting (2018, 2019). What do you plan to do during the next reporting period to accomplish the goals?Unfortunately, though this project was a useful experience in training the student in research methodology, trouble-shooting, and critical thinking, there is little that can be done at this point to meet the original research objectives as the samples have all been expended. Given the nature of the multigenerational study, generating new samples is not feasible.
Impacts
What was accomplished under these goals?
Originally, we had proposed to execute the proposed work using the Qiagen DNA Stool Mini, followed by sequencing on the Iontorrent. Unfortunately both of these options were discontinued before the study had been funded. We now have the Illumina Nextseq platform available in our Core Facilities at Utah State University. Utilization of the Illumina Nextseq will allow me to multiplex all of my samples into one run, thereby decreasing variability that is typically experienced run-to-run. Much of the grant period was focused on developing and optimizing protocols for fecal sample processing using alternative kits and protocols for downstream sequencing on the new platform. We established a dual primer set for multiplexing on this instrument, as well as an enhanced V3 primer sequence for increased coverage rate. Several commercially available kits were evaluated for use for our particular samples, and the DNeasy PowerSoil Kit was identified as the best choice for this work with a modified protocol for cleaning DNA samples.
Publications
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2019
Citation:
Presentations
Bartlett, A. (Presenter & Author), Faslev, K., Catalano, L. (Author Only), Childress, T. (Author Only), Perry, J. (Author Only), Buhler, J. (Author Only), Child, N. (Author Only), Dygert, K. (Author Only), Phatak, S. (Author Only), Hintze, K. J. (Author Only), Benninghoff, A. (Author Only), Research Week: Student Research Symposium, "Basal diet, green tea extract and gut microbiome interactions in a mouse multi-generation study.," Utah State University, Logan, UT. (April 8, 2019 - April 12, 2019)
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2019
Citation:
Presentations
Phatak, S. (Presenter & Author), Thomas, A. (Author Only), Kaundal, R. (Author Only), Jones, R. (Author Only), Hintze, K. J., Benninghoff, A. (Author Only), Research Week: Student Research Symposium, "Impact of the total Western diet for rodents on colon mucosal gene expression in a multigenerational murine model of colitis-associated colorectal cancer," Utah State University, Logan, UT. (April 8, 2019 - April 12, 2019)
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Progress 03/15/18 to 03/14/19
Outputs
Target Audience:Research scientists Changes/Problems:The major challenges in the past year were rooted in having to change sequencing platforms and the discontinuation of the kit we use for isolating DNA. Those have largely been addressed at this point, and no further changes or major problems are expected. What opportunities for training and professional development has the project provided?During the past year, Sumira has participated in many professional development events and other training opportunities. For example, she has routinely attended the Graduate Research Training Series at USU; a lecture on rethinking science communication; workshops on resume's/cover letters and job negotiations, and other relevant webinars and colloquiums. Of special note, Sumira competed for and won a slot at a week-long study abroad course focused on the Mediterranean Diet. How have the results been disseminated to communities of interest?The microbiome data are not yet available for presentation. However, Sumira has been presenting her overall dissertation research at several conferences including the Environmental Mutagenesis Society (2018), the American Society for Nutrition annual meeting (2018) and the Society of Toxicology annual meeting (2019). What do you plan to do during the next reporting period to accomplish the goals?With an optimized, working protocol for the new sequencing platform, Sumira will complete the ongoing extractions of fecal DNA for sequencing, submit samples for sequencing, and complete data analysis in addition to her other dissertation work.
Impacts
What was accomplished under these goals?
Originally, we had proposed to execute the proposed work using the Qiagen DNA Stool Mini, followed by sequencing on the Iontorrent. Unfortunately both of these options were discontinued before the study had been funded. We now have the Illumina Nextseq platform available in our Core Facilities at Utah State University. Utilization of the Illumina Nextseq will allow me to multiplex all of my samples into one run, thereby decreasing variability that is typically experienced run-to-run. Much of the past year has been focused on developing and optimizing protocols for fecal sample processing using a alternative kits and protocols for downstream sequencing on the new platform. We have established a dual primer set for multiplexing on this instrument, as well as an enhanced V3 primer sequence for increased coverage rate. Several commercially available kits were evaluated for use for our particular samples, and the DNeasy PowerSoil Kit was identified as the best choice for this work.
Publications
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