3 research outputs found
A new way to rapidly create functional, fluorescent fusion proteins: random insertion of GFP with an in vitro transposition reaction
BACKGROUND: The jellyfish green fluorescent protein (GFP) can be inserted into the middle of another protein to produce a functional, fluorescent fusion protein. Finding permissive sites for insertion, however, can be difficult. Here we describe a transposon-based approach for rapidly creating libraries of GFP fusion proteins. RESULTS: We tested our approach on the glutamate receptor subunit, GluR1, and the G protein subunit, α(s). All of the in-frame GFP insertions produced a fluorescent protein, consistent with the idea that GFP will fold and form a fluorophore when inserted into virtually any domain of another protein. Some of the proteins retained their signaling function, and the random nature of the transposition process revealed permissive sites for insertion that would not have been predicted on the basis of structural or functional models of how that protein works. CONCLUSION: This technique should greatly speed the discovery of functional fusion proteins, genetically encodable sensors, and optimized fluorescence resonance energy transfer pairs
The microbiome of deep-sea fish reveals new microbial species and a sparsity of antibiotic resistance genes
Adaptation to life in the deep-sea can be dramatic, with fish displaying behaviors and appearances
unlike those seen in any other aquatic habitat. However, the extent of which adaptations may have
developed at a microbial scale is not as clear. Shotgun metagenomic sequencing of the intestinal
microbiome of 32 species of deep-sea fish from across the Atlantic Ocean revealed that many of the
associated microbes differ extensively from those previously identified in reference databases. 111
individual metagenome-assembled genomes (MAGs) were constructed representing individual
microbial species from the microbiomes of these fish, many of which are potentially novel bacterial
taxa and provide a window into the microbial diversity in this underexplored environment. These
MAGs also demonstrate how these microbes have adapted to deep-sea life by encoding a greater
capacity for several cellular processes such as protein folding and DNA replication that can be
inhibited by high pressure. Another intriguing feature was the almost complete lack of genes
responsible for acquired resistance to known antibiotics in many of the samples. This highlights that
deep-sea fish microbiomes may represent one of few animal-associated microbiomes with little
influence from human activity. The ability of the microbes in these samples to bioluminesce is lower
than expected given predictions that this trait has an important role in their life cycle at these
depths. The study highlights the uniqueness, complexity and adaptation of microbial communities
living in one of the largest and harshest environments on Earth