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SilflowCD

Research Abstracts Online
January - December 2011

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University of Minnesota Twin Cities
College of Biological Sciences
College of Food, Agricultural, and Natural Resource Sciences
Department of Plant Biology

PI: Carolyn D. Silflow

Use of Next-Generation Sequencing in the Analysis of Genes in the Hydrogen Biosynthesis Pathway in Chlamydomonas

Under anaerobic conditions, the green alga Chlamydomonas reinhardtii produces hydrogen gas in a reaction catalyzed by hydrogenase. The expression of two hydrogenase genes, HYDA1 and HYDA2, is strictly regulated by anaerobiosis, through mechanisms that are still unknown. To identify trans-acting genes in this pathway, these researchers have designed a mutant screen using a motility gene reporter driven by a HYDA1 promoter. The screen of chemically mutagenized cells identified a mutant strain B6-F that, under aerated conditions, is motile and expresses HYDA1 and other hydrogenase-related genes at elevated levels. The mutant phenotype is recessive in stable heterozygous diploids. Genetic mapping placed this mutation to a ~ 1 Mbp region on chromosome 1. Analysis of Illumina next-generation sequencing data revealed the presence of an extensive number of SNPs between the genomes of mutant strain B6-F and the strain from which it was derived, B6. Within the 1Mbp region, the researchers identified one SNP mutation that disrupted a splicing signal in an unannotated gene encoding a protein of 52 kD. The incorrectly spliced transcript in the mutant strain has been verified by RT-PCR. The cloned wild-type gene is being tested for genetic rescue to further examine the relationship between the mutation and the elevated expression level of HYDA1 and hydrogenase-related genes. Data from transcriptome sequencing will be used to elucidate the role of the newly identified gene in the signaling pathway leading to transcription of the hydrogenase genes.

Group Member

Xiaoqing Sun, Graduate Student