The increasing ability to sequence and compare multiple individual genomes within a species has highlighted the fact that copy-number variation (CNV) is a substantial and underappreciated source of genetic diversity. Chromosome-scale mutations occur at rates orders of magnitude higher than base substitutions, yet our understanding of the mechanisms leading to CNVs has been lagging. We examined CNV in a region of chromosome 5 (chr5) in haploid and diploid strains of Saccharomyces cerevisiae. We optimized a CNV detection assay based on a reporter cassette containing the SFA1 and CUP1 genes that confer gene dosage-dependent tolerance to formaldehyde and copper, respectively. This optimized reporter allowed the selection of low-order gene amplification events, going from one copy to two copies in haploids and from two to three copies in diploids. In haploid strains, most events involved tandem segmental duplications mediated by nonallelic homologous recombination between flanking direct repeats, primarily Ty1 elements. In diploids, most events involved the formation of a recurrent nonreciprocal translocation between a chr5 Ty1 element and another Ty1 repeat on chr13. In addition to amplification events, a subset of clones displaying elevated resistance to formaldehyde had point mutations within the SFA1 coding sequence. These mutations were all dominant and are proposed to result in hyperactive forms of the formaldehyde dehydrogenase enzyme.A S a consequence of studies that utilize genomic microarrays and next-generation DNA sequencing, it has become clear that much of the natural genetic variation that exists between individuals is due to alterations in the number of copies of genes rather than differences in the nucleotide sequence (Girirajan et al. 2011;Veltman and Brunner 2012). It has been calculated that 8-25 kb of DNA are deleted or duplicated per generation in humans compared to about 100 bp of point mutations (Itsara et al. 2010). Although deletions and duplications vary in size from a few base pairs (for example, changes in the lengths of microsatellites) to many megabases (for example, changes in ploidy), Girirajan et al. (2011) define copy-number variants (CNVs) in humans as changes that vary in size between 50 bp and 1 Mb. While most CNV events have no obvious effect, a significant number are associated with human diseases including Charcot-Marie-Tooth syndrome, autosomal dominant leukodystrophy, Williams syndrome, several cancer predispositions, and autism-related and other neurodevelopmental disorders (Abrahams and Geschwind 2008;Girirajan et al. 2011;Krepischi et al. 2012;Malhotra and Sebat 2012;Sullivan et al. 2012). There is also strong evidence suggesting that CNVs played a significant role in human evolution (Iskow et al. 2012). Finally, multiple somatic CNV events are frequently observed in the altered karyotypes of cancer cells (Stratton et al. 2009).CNVs have also been widely observed in natural populations and have been studied in detail in model organisms. In the discussion below, w...
BackgroundSynthetic biology allows the development of new biochemical pathways for the production of chemicals from renewable sources. One major challenge is the identification of suitable microorganisms to hold these pathways with sufficient robustness and high yield. In this work we analyzed the genome of the propionic acid producer Actinobacteria Propionibacterium acidipropionici (ATCC 4875).ResultsThe assembled P. acidipropionici genome has 3,656,170 base pairs (bp) with 68.8% G + C content and a low-copy plasmid of 6,868 bp. We identified 3,336 protein coding genes, approximately 1000 more than P. freudenreichii and P. acnes, with an increase in the number of genes putatively involved in maintenance of genome integrity, as well as the presence of an invertase and genes putatively involved in carbon catabolite repression. In addition, we made an experimental confirmation of the ability of P. acidipropionici to fix CO2, but no phosphoenolpyruvate carboxylase coding gene was found in the genome. Instead, we identified the pyruvate carboxylase gene and confirmed the presence of the corresponding enzyme in proteome analysis as a potential candidate for this activity. Similarly, the phosphate acetyltransferase and acetate kinase genes, which are considered responsible for acetate formation, were not present in the genome. In P. acidipropionici, a similar function seems to be performed by an ADP forming acetate-CoA ligase gene and its corresponding enzyme was confirmed in the proteome analysis.ConclusionsOur data shows that P. acidipropionici has several of the desired features that are required to become a platform for the production of chemical commodities: multiple pathways for efficient feedstock utilization, ability to fix CO2, robustness, and efficient production of propionic acid, a potential precursor for valuable 3-carbon compounds.
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