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United States Department of Agriculture

Agricultural Research Service

Research Project: GENETIC DISSECTION OF SEEDLING COLD TOLERANCE IN RICE (ORYZA SATIVA L.)

Location: Crops Pathology and Genetics Research

Title: Reference genome-independent assessment of mutation density using restriction enzyme-phased sequencing

Authors
item Monson-Miller, Jennifer -
item Sanchez-Mendez, Diana -
item TAI, THOMAS
item Comai, Luca -

Submitted to: Biomed Central (BMC) Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: February 14, 2012
Publication Date: February 17, 2012
Repository URL: http://www.biomedcentral.com/1471-2164/13/72
Citation: Monson-Miller, J., Sanchez-Mendez, D.C., Tai, T., Comai, L. 2012. Reference genome-independent assessment of mutation density using restriction enzyme-phased sequencing. Biomed Central (BMC) Genomics. 13:72.

Interpretive Summary: Inducing mutations using chemical or physical agents (i.e. mutagens) is a powerful approach to generating genetic variation for gene discovery as well as germplasm enhancement. One important goal is determining the effectiveness of a mutagenic treatment (i.e. the mutation density or rate) prior to further investment in time and resources in characterizing the treated materials. Since the frequency of visible mutations does not always reflect the actual mutation density, direct analysis of the DNA by sequencing is the best approach. Here we report the development and application of a simple, rapid, and cost-effective method to assess mutation density which can be used in the absence of reference genome sequence. Thus, this method can be employed to identify mutant populations with high mutation densities to improve the efficiency of downstream analyses and can be applied in species whose genomes have not yet been sequenced.

Technical Abstract: Determining the mutation rates in single individuals is a significant challenge requiring a substantial amount of sequencing and analysis. We have developed a simple method based on restriction enzyme-phased sequencing to measure the mutation density by comparing a wild type to mutagenized individuals. Another advantage of this method is its independence from the availability of reference genome sequence. We demonstrate the efficacy of this approach using rice.

Last Modified: 9/10/2014