PubMed · 8257953
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J H Beijnen. 1993-10-15. Does antisense make sense?. https://doi.org/10.1007/bf01880623
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STALARD (selective target amplification for low-abundance RNA detection) enables isoform-level quantification of low-abundance RNAs using conventional laboratory equipment. Here, we describe steps for RNA isolation, primer design, reverse transcription, selective target amplification, and downstream analysis. The protocol couples selective pre-amplification with a quantitative reverse-transcription PCR (RT-qPCR) readout and optional nanopore sequencing. Using 1 μg input RNA and 12 pre-amplification cycles, STALARD reduces Cq values by approximately 10-12 cycles, bringing the target into a reliably quantifiable range. For complete details on the use and execution of this protocol, please refer to Jeong et al.1.
The expression of a key mitochondrial membrane component, the ADP/ATP carrier, was investigated in two aerobic yeast species, Kluyveromyces lactis and Schizosaccharomyces pombe. Although the two species differ very much in their respiratory capacity, the expression of the carrier in both yeast species was decreased under partially anaerobic conditions and was induced by nonfermentable carbon sources. The single ADP/ATP carrier encoding gene was deleted in S. pombe. The null mutant exhibits impaired growth properties, especially when cultivated at reduced oxygen tension, and is unable to grow on a nonfermentable carbon source. Our results suggest that the inability of K. lactis and S. pombe to grow under anaerobic conditions can be related in part to the absence of a functional ADP/ATP carrier due to repression of the corresponding gene expression.
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