DNA pairing is an important step in the process of targeted nucleotide exchange

Modified single-stranded DNA oligonucleotides can direct the repair of genetic mutations in yeast, plant and mammalian cells. The mechanism by which these molecules exert their effect is being elucidated, but the first phase is likely to involve the homologous alignment of the single strand with its complementary sequence in the target gene. In this study, we establish the importance of such DNA pairing in facilitating the gene repair event. Oligonucleotide-directed repair occurs at a low frequency in an Escherichia coli strain (DH10B) lacking the RECA DNA pairing function. Repair activity can be rescued by using purified RecA protein to catalyze the assimilation of oligonucleotide vectors into a plasmid containing a mutant kanamycin resistance gene in vitro. Electroporation of the preformed complex into DH10B cells results in high levels of gene repair activity, evidenced by the appearance of kanamycin-resistant colonies. Gene repair is dependent on the formation of a double-displacement loop (double-D-loop), a recombination intermediate containing two single-stranded oligonucleotides hybridized to opposite strands of the plasmid at the site of the point mutation. The heightened level of stability of the double-D-loop enables it to serve as an active template for the DNA repair events. The data establish DNA pairing and the formation of the double-D-loop as important first steps in the process of gene repair

Marine Population Genomics: Challenges and Opportunities

Population genomics is revolutionizing biology and stimulating new research questions and directions. While human health has driven many of the genomics tools and approaches, all other biological fields have benefitted. This is certainly true in the world’s oceans, which encompass a large diversity of species and ecosystems. In the world’s oceans, population genomics approaches are giving us an unprecedented ability to gain a better understanding of the organisms inhabiting these ecosystems. While population genomics approaches are improving our understanding of genetic diversity and population genetic parameters in marine organisms, they also are providing unexpected insights into marine invasions, population connectivity, and how marine organisms are responding to different stimuli and environments. Some examples include identification of connectivity among populations that is not predicted by geography as well as identification of genes and genetic variants under natural selection in response to environment and climate conditions as indicators of genes and pathways responsible for adaption. This knowledge is important because so much of the world’s oceans is understudied. This knowledge also is critical for understanding how marine organisms will respond to environmental change and thus how we can better protect marine biodiversity and marine resources. That is, we can better predict the effects of enhanced migration on mitigating anthropogenic stressors affecting marine populations and whether outcrossing will enhance population survival or result in outbreeding depression. Simply put, population genomics provides the genetic resolution to make better predictions about how environmental change is affecting populations and thus provides insights into how we might address environmental change’s deleterious effects on important marine resources. In this chapter, we provide an overview of the challenges and opportunities for marine population genomics, addressing how population genomics can be used to understand marine biodiversity, population demographics and connectivity, and response to environmental changes as well as assist sustainable management, protection, and conservation of marine biodiversity