News, Selected Publications & Resources

Large-Scale Reverse Genetics in Arabidopsis: Case Studies from the Chloroplast 2010 Project

Article link: Plant Phys 152:529 (2010)
Researchers at Michigan State University took part in the Chloroplast 2010 Project, designed to screen thousands of sequence-indexed Arabidopsis thaliana T-DNA insertion mutants for various phenotypes. In order to have confidence that a given mutant phenotype is associated with an alteration to a specific gene, the team looked for two or more independent homozygous mutants for each target gene. A high-throughput method was essential for this number of queries. The team employed a first-pass SYBR® Green method to eliminate homozygous samples. The remaining samples were subjected to further analysis to distinguish between heterozygous and wild-type genotypes using the two-reaction method and agarose gel electrophoresis involving Applied Biosystems® SYBR® Green PCR Master Mix and AmpliTaq Gold® with PCR on the 7900 HT Sequence Detection System.

Sulfur Transfer through an Arbuscular Mycorrhiza

Article link: Plant Phys 149:549 (2008)
In an investigation aimed at the role of arbuscular mycorrhizal symbiosis in sulfur (S) uptake in plants was undertaken by a team at Michigan State University, USA. The researchers sought to determine which forms of S were taken up, metabolized, and transferred to the plant host roots by the fungal partner and to identify fungal genes were responsible for handling S. To examine fungal genes potentially involved in S transport, total RNA was isolated from frozen germinating spore tissue, and cDNA was synthesized using SuperScript® II reverse transcriptase. Gene fragments were generated using PCR and gene-specific primers, and were sequenced using an Applied Biosystems® 3130 Genetic Analyzer. Real-time PCR (using the Applied Biosystems® 7900 HT Sequence Detection System and Power SYBR® Green 2-Step Master Mix) revealed changes in a gene involved in sulfate transport and assimilation, which indicates a role for transcriptional regulation of sulfate permease in the reduction in sulfate transport in the presence of cysteine.