Abstract
The hybridization of mitochondrial leucyl- (mit-leucyl-) tRNA and mit-valyl-tRNA with mitochondrial and nuclear DNA of grande yeast has been studied. The tRNA's were charged in vitro with amino acids of high specific radioactivity and hybridization carried out at 33 °C in formamide at pH 5.0 to minimize deacylation of the tRNA. The mit[ 3H]leucyl- and mit[ 3H]valyl-tRNA's hybridized with mitochondrial DNA but not with yeast nuclear DNA or Escherichia coli DNA. Apparent hybridization-saturation curves were obtained. The melting profile of the hybrid was sharp and equivalent to a T m of 71 °C in 2 × standard saline citrate. Unlabeled yeast mitochondrial tRNA competed for the hybridization, but yeast supernatant and E. coli tRNA's did not. RNase and alkaline digests of the mit[ 3H]valyl-tRNA-mit-DNA hybrid yielded [ 3H]valyladenosine and [ 3H]valine, respectively, providing further evidence for the validity of the hybridization system.
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