Identifying mutations in Arabidopsis – a faster, cheaper method


Highlighted paper: Liu, McCormack and Sheen (2012) Targeted parallel sequencing of large genonmic regions for identifying mutations in Arabidopsis. Plant Methods 8:12

Kun-hsiang Liu, Matthew McCormack and Jen Sheen from Harvard have developed a PCR-based method of identifying mutations in Arabidopsis. It is cheaper and easier than traditional methods of identifying mutations, using bench-top PCR and a new user-friendly method of bioinformatics analysis using web-based resource Galaxy. Liu et al. estimate that using their method to identify a mutation mapped to a 550kb genomic region will cost roughly US$500, a fraction of the usual ten thousand dollar cost of currently used methods of mutant identification.

Liu et al. tested the new method of identifying mutations by searching for new nitrogen response genes. They generated an Arabidopsis thaliana line in which LUCIFERASE was driven by the promoter for nitrogen response marker NIR. Using EMS-mutagenesis, the team made 25 000 mutant NIR:LUC lines and identified seedlings that were nitrate insensitive (nis) or showed nitrate constitutive response (ncr).

When the lines were made, the phenotypes were identified and a second generation was grown. Three second generation lines – ncr1, nis1 and nis2 – were selected for further investigation.

Liu et al. used their novel TPSeq method to locate the mutations causing the ncr1, nis1 and nis2 phenotypes. (more…)

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