Family-specific vs. universal PCR primers for the study of mitochondrial DNA in plants

Abstract

Mitochondrial genomes (mtDNAs or mitogenomes) of seed plants are characterized by a notoriously unstable organization on account of which available so-called universal or consensus primers may fail to fulfil their foreseen function-amplification of various mtDNA regions in a broad range of plant taxa. Thus, the primers developed for groups assumed to have similar organization of their mitogenomes, such as families, may facilitate a broader usage of more variable non-coding portions of these genomes in group members. Using in silico PCR method and six available complete mitogenomes of Fabaceae, it has been demonstrated that only three out of 36 published universal primer and three Medicago sativa-specific primer pairs that amplify various mtDNA regions are suitable for six representatives of the Fabaceae family upon minor modifications, and develop 21 Fabaceae-specific primer pairs for amplification of all 14 cis-splicing introns in genes of NADH subunits (nad genes) which represent the most commonly used noncoding mtDNA regions in various studies in plants. Using the same method and six available complete mitogenomes of representatives of related families Cucurbitaceae, Euphorbiaceae and Rosaceae and a model plant, Arabidopsis thaliana, it has further been demonstrated that applicability of newly developed primer pairs for amplification of nad introns in more or less related taxa was dependent not only on species evolutionary distances but also on their genome sizes. A reported set of 24 primer pairs is a valuable resource which may facilitate a broader usage of mtDNA variability in future studies at

Plant mitochondrial introns as genetic markers - conservation and variation

“…This expands and improves the available universal primers for plant mitochondrial introns ( Demesure et al., 1995 ; Dumolin-Lapegue et al., 1997 ; Duminil et al., 2002 ). Aleksić (2016) found limited applicability of previously developed universal mitochondrial primers to legume ( Fabaceae ) species and suggested family-specific primers as a more practical approach. The primer sets employed here successfully amplified the mitochondrial introns of P. vulgaris as a representative legume.…”