Abstract
BackgroundPerennial ryegrass (Lolium perenne L.) is one of the most important forage and turf grass species of temperate regions worldwide. Its mitochondrial genome is inherited maternally and contains genes that can influence traits of agricultural importance. Moreover, the DNA sequence of mitochondrial genomes has been established and compared for a large number of species in order to characterize evolutionary relationships. Therefore, it is crucial to understand the organization of the mitochondrial genome and how it varies between and within species. Here, we report the first de novo assembly and annotation of the complete mitochondrial genome from perennial ryegrass.ResultsIntact mitochondria from perennial ryegrass leaves were isolated and used for mtDNA extraction. The mitochondrial genome was sequenced to a 167-fold coverage using the Roche 454 GS-FLX Titanium platform, and assembled into a circular master molecule of 678,580 bp. A total of 34 proteins, 14 tRNAs and 3 rRNAs are encoded by the mitochondrial genome, giving a total gene space of 48,723 bp (7.2%). Moreover, we identified 149 open reading frames larger than 300 bp and covering 67,410 bp (9.93%), 250 SSRs, 29 tandem repeats, 5 pairs of large repeats, and 96 pairs of short inverted repeats. The genes encoding subunits of the respiratory complexes – nad1 to nad9, cob, cox1 to cox3 and atp1 to atp9 – all showed high expression levels both in absolute numbers and after normalization.ConclusionsThe circular master molecule of the mitochondrial genome from perennial ryegrass presented here constitutes an important tool for future attempts to compare mitochondrial genomes within and between grass species. Our results also demonstrate that mitochondria of perennial ryegrass contain genes crucial for energy production that are well conserved in the mitochondrial genome of monocotyledonous species. The expression analysis gave us first insights into the transcriptome of these mitochondrial genes in perennial ryegrass.
Highlights
Perennial ryegrass (Lolium perenne L.) is one of the most important forage and turf grass species of temperate regions worldwide
Isolation of intact mitochondria and extraction of mitochondrial DNA (mtDNA) A cellular fraction containing crude mitochondria were isolated from perennial ryegrass leaf tissue by homogenization followed by differential centrifugation
The crude mitochondrial fraction was characterized by measuring the activity and latency of cytochrome c oxidase (CCO) as a marker enzyme for the intactness of the inner mitochondrial membrane, and malate dehydrogenase (MDH), an enzyme residing in the mitochondrial matrix as well as in the cytosol and several other places in the cell [13] (Table 1)
Summary
Perennial ryegrass (Lolium perenne L.) is one of the most important forage and turf grass species of temperate regions worldwide. Their primary function is the production of metabolic intermediates and cellular ATP through the citric acid cycle and oxidative phosphorylation pathway. For this reason, mitochondria are involved in a wide variety of cellular and developmental processes including pollen development and cytoplasmic male sterility (CMS) [1,2]. Mitochondria are involved in a wide variety of cellular and developmental processes including pollen development and cytoplasmic male sterility (CMS) [1,2] Mitochondria have their own genomes, which harbor variation in both size and actual gene content, despite the universally conserved sequence that exists between the mitochondrial genomes of diverse species [3]. Sequencing of the mitochondrial genome has the potential to increase our understanding of the complex genetic interactions between the nuclear and the organellar genomes
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