CC-4871 rbcL-K175S mt+
$30.00
From Robert J. Spreitzer, University of Nebraska, August 2014
Phenotype: requires acetate, sensitive to light
Using plasmid p699 (GuhaMajumdar et al. 2008), Sriram Satagopan in Spreitzer’s group performed rbcL direct mutagenesis, chloroplast transformation of wild-type 2137 mt+ (cloned), selection for spectinomycin resistance in the dark, and screening for a homoplasmic acetate-requiring phenotype to create a K175G substitution (AAA-TCA) in the Rubisco large subunit. The K175S substitution causes a 90% decrease in Rubisco holoenzyme and a 99% decrease in Rubisco carboxylase activity (Lim and Spreitzer, unpublished). This mutant was created to investigate the role of active-site Lys-175 in catalysis (Harpel et al. 2002). It has been maintained with acetate medium in darkness since its creation.
GuhaMajumdar M, Dawson-Baglien E, Sears BB (2008) Creation of a chloroplast microsatellite reporter for detection of replication slippage in Chlamydomonas reinhardtii. Eukaryot Cell 7:639-646
Harpel MR, Larimer FW, Hartman FC (2002) Multifaceted roles of Lys166 of ribulose-bisphosphate carboxylase/oxygenase as discerned by product analysis and chemical rescue of site-directed mutants. Biochemistry 41:1390-1397