| Publication Type |
thesis |
| School or College |
School of Medicine |
| Department |
Pathology |
| Author |
Seipp, Michael Todd |
| Title |
Genotyping by high-resolution amplicon melting |
| Date |
2010-01-13 |
| Description |
Genotyping by high-resolution amplicon melting is a simple method using only PCR primers and double-stranded DNA binding dyes. Heterozygous genotypes are easily assigned based on derivative melting curve shape and width. Homozygous genotypes are assigned based on Tm differences that are usually 1 °C but can be less. One potential drawback to genotyping by amplicon melting is Tm variance that can be caused by solution chemistry differences and thermal variability of PCR instruments. To address the Tm variance, internal temperature standards were created using complimentary 3'-blocked oligonucleotides which were included in each reaction. Genotyping by amplicon melting lends itself to potential multiplexing, due to only primers being used for detection. A thrombophilia panel, including the F5, F2, and the MTHFR 677 and 1298 mutations, was used as a multiplex model because of the potential additive effects of mutations at multiple loci. The first set of experiments were done using the LightCycler 1.5 and HR-1 technologies, which performed well in a blinded study of 109 samples. This assay was further improved by conversion to a mutli-sample high-resolution platform combining the LightCycler and HR-1 technologies. This new process decreased hands-on manipulation as well as overall analysis time while maintaining the same resolution. |
| Type |
Text |
| Publisher |
University of Utah |
| Subject |
Genomics; Gene Amplification |
| Subject MESH |
Genomics; Gene Amplification |
| Dissertation Institution |
University of Utah |
| Dissertation Name |
MS |
| Language |
eng |
| Relation is Version of |
Digital reproduction of "Genotyping by high-resolution amplicon melting." Spencer S. Eccles Health Sciences Library. Print version of "Genotyping by high-resolution amplicon melting." available at J. Willard Marriott Library Special Collection. QH9.7 2009.S45 |
| Rights Management |
© Michael Todd Seipp |
| Format Medium |
application/pdf |
| Format Extent |
849,418 bytes |
| Source |
Original: University of Utah Spencer S. Eccles Health Sciences Library |
| Conversion Specifications |
Original scanned on Fujitsu fi-5220G as 400 dpi to pdf using ABBYY FinerReader 10 |
| ARK |
ark:/87278/s65d96dg |
| Setname |
ir_etd |
| ID |
193290 |
| Reference URL |
https://collections.lib.utah.edu/ark:/87278/s65d96dg |
