Undergraduate Thesis Or Project
 

Sampson_BRR_Thesis.pdf

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https://ir.library.oregonstate.edu/concern/undergraduate_thesis_or_projects/nk322g21d

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  • Lung cancer in humans is an adult-onset disease. It is the number one cause of cancer mortality for both men and women and has a poor (15%) survival rate. Biological markers of the disease would allow for early diagnosis and treatment. The specific aims of this research project were to develop quality control and miRNA isolation methods to prepare lung tissues of newborn and adult mice exposed to lung carcinogens in utero for miRNA sequencing. A quality analysis with biological and technical replicates was completed on the Illumina HiSeq 2000 platform. The method of RNA extraction needed to provide sufficient yield from small pieces of fibrous tissue (low cell density) for downstream applications. Three RNA isolation kits and a proprietary phenol-chloroform extraction method were chosen to test for feasibility in this particular application, as were two homogenizing methods. The three kits (all spin column-based extraction methods) and proprietary method included: Roche High Pure miRNA Isolation Kit, Qiagen RNeasy Plus Micro Kit, Qiagen RNeasy Mini Kit, and Trizol. Frosted Pyrex homogenizers were selected over the Tissue Tearor due to the size and type of tissue being homogenized. The spin column-based methods produced high quality total RNA, but yielded amounts too low to sequence. The Trizol extraction method was able to yield large amounts of high quality RNA and preserved the small RNAs. Sequencing data from technical and biological replicates were compared (R² values between 0.995 and 0.999) and found to be highly reproducible. These data confirm the reliability and reproducibility of miRNA sequencing on the Illumina 2000 sequencer.
  • Keywords: dibenzo[def, p]chrysene, miRNA, lung cancer
  • Keywords: dibenzo[def, p]chrysene, miRNA, lung cancer
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file details Sampson_BRR_Thesis.pdf 2017-07-27 Pubblico Scaricare