Simultaneous Identification of Multiple Causal Mutations in Rice.

Authors:
Wei Yan
Wei Yan
South China Normal University
PhD
Molecular crop breeding
Guangzhou, Guangdong | China
Jiawei Lu
Jiawei Lu
Soochow University
China
Chunjue Xu
Chunjue Xu
National Key Laboratory of Crop Genetic Improvement and National Center of Plant Gene Research (Wuhan)
Gang Xie
Gang Xie
Key Laboratory of Synthetic and Natural Functional Molecule Chemistry of Ministry of Education
United States
Yiqi Li
Yiqi Li
Fujian University of Traditional Chinese Medicine
Xing Wang Deng
Xing Wang Deng
Peking University
China
Hang He
Hang He
Peking University
China

Front Plant Sci 2016 17;7:2055. Epub 2017 Jan 17.

Shenzhen Institute of Molecular Crop DesignShenzhen, China; Guangdong Provincial Key Laboratory of Biotechnology for Plant Development, College of Life Sciences, South China Normal UniversityGuangzhou, China.

Next-generation sequencing technologies (NGST) are being used to discover causal mutations in ethyl methanesulfonate (EMS)-mutagenized plant populations. However, the published protocols often deliver too many candidate sites and sometimes fail to find the mutant gene of interest. Accurate identification of the causal mutation from massive background polymorphisms and sequencing deficiencies remains challenging. Here we describe a NGST-based method, named SIMM, that can simultaneously identify the causal mutations in multiple independent mutants. Multiple rice mutants derived from the same parental line were back-crossed, and for each mutant, the derived F2 individuals of the recessive mutant phenotype were pooled and sequenced. The resulting sequences were aligned to the Nipponbare reference genome, and single nucleotide polymorphisms (SNPs) were subsequently compared among the mutants. Allele index (AI) and Euclidean distance (ED) were incorporated into the analysis to reduce noises caused by background polymorphisms and re-sequencing errors. Corrections of sequence bias against GC- and AT-rich sequences in the candidate region were conducted when necessary. Using this method, we successfully identified seven new mutant alleles from Huanghuazhan (HHZ), an elite rice cultivar in China. All mutant alleles were validated by phenotype association assay. A pipeline based on Perl scripts for SIMM is publicly available at https://sourceforge.net/projects/simm/.

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Source
http://dx.doi.org/10.3389/fpls.2016.02055DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5239786PMC
January 2017
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