Abstract 2095: A virtual pyrogram generator to resolve ambiguity of pyrosequencing results
| Autor: | Matthew T. Olson, Alan O'Neill, Marija Debeljak, Katie Beierl, Shuko Harada, Ming-Tseh Lin, Christopher D. Gocke, Keila Rivera-Roman, Alexis Norris-Kirby, Samantha Finley, James R. Eshleman, Amanda Stafford, Guoli Chen |
|---|---|
| Rok vydání: | 2012 |
| Předmět: | |
| Zdroj: | Cancer Research. 72:2095-2095 |
| ISSN: | 1538-7445 0008-5472 |
| DOI: | 10.1158/1538-7445.am2012-2095 |
| Popis: | We report a freely available software program, Pyromaker, which generates simulated pyrosequencing traces based on user inputs and aids in analysis of complex pyrosequencing results. To achieve this goal, simulated pyrograms are generated based on the hypothesized mutations and matched with the actual pyrogram. We validated the software testing common KRAS gene mutations and several mutations in the BRAF, GNAS and, p53 genes. We demonstrate that all eighteen possible single base mutations within codons 12 and 13 of KRAS gene generate unique simulated pyrosequencing traces and highlight the distinctions between them. We further verify that all reported codon 12 and 13 complex mutations produce unique pyrograms. However, some complex pyrograms are indistinguishable from single base mutations, using a standard dispensation order. Pyromaker was used in two strategies, one in which hypothesis-based simulated pyrograms were pattern-matched with the actual pyrograms. In a second mode, with only the pyrogram, Pyromaker was used to identify the underlying mutation by iteratively reconstructing the mutant pyrogram. Either strategy was able to successfully identify the complex mutations, which were confirmed by cloning and subsequent sequencing. With two examples of KRAS codon 12 mutations, (GGTαTTT, G12F and GGTαGAG, G12E), we report optimal combinations of the five analysis approaches that permit unambiguous mutation identification. The most efficient one is pyrosequencing with Pyromaker. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2095. doi:1538-7445.AM2012-2095 |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|