-
This group of microorganisms constitutes one of the three domains (major branches) of life. Discovered by Carl Woese in the late 1970's, archaea were originally thought to be those organisms that lived only in extreme environments. These organisms share many common biological features that set them aside from the rest of the bacteria that microbiologists had studied for years. For instance, their outer membranes are made of ether-linked rather than ester-linked fatty acids like the rest of biological life, and the way that they process their DNA and RNA in the cell is much more like a eukaryote. By comparing gene features that are common to all forms of life, in this case the ribosomal RNA, Dr. Woese found that the extreme environment loving microorganisms appeared to share a common evolutionarily origin - and that they were sufficiently different from the bacteria and eukaryotes that he proposed a third form (Domain) of life, the Archaebacteria. Since then the name has been shortened to Archaea, and these organisms once thought to only inhabit extreme environments are found in nearly every environment that we look! They are the dominant life form in many extreme environments, but the group is much more diverse that originally recognized, such that relatives of the hyperthermophiles (organisms living at temperatures > 80) have been found in seawater, lakes, and soils throughout the world.
- bacterioplankton
- Organisms living in aquatic habitats are classified by size (megaplankton, mesoplankton, microplankton, nanoplankton, picoplankton) and biological group (zooplankton, phytoplankton, bacterioplankton). The bacterioplankton encompass both bacteria and archaea that are free-living in aquatic habitats.
-
Metabolites, or chemicals that microbes produce during their daily life functions. These can include proteins/enzymes, ribonucleic acids, fatty acids, antibiotics, gases, minerals, and other chemicals. Detection of these metabolites and chemicals can provide clues as to which processes the organisms are involved with, which chemical transformations they are performing, and how they respond to changes in their environment.
-
Denaturing Gradient Gel Electrophoresis - a molecular biology tool used to separate DNA fragments by their sequence specific melting (denaturation) point. DGGE has increasingly been used to visualize the diversity of ribosomal RNA genes present in natural microbial samples from all sorts of habitats. After a sample is run through the gel, individual sequences, representing different species show up as distinct bands - therefore providing a fingerprint of the diversity in a sample. You can run 15-20 samples on a gel at a time, and compare the fingerprints within and between gels.
-
Organisms that thrive in niche environments where most organisms are not adapted for survival. Such environments include but are not limited to sulfur springs (lacking oxygen), thermal vents (heat extremes), brine channels (extremes in salt), and polar environments (cold extremes).
-
An f-factor cosmid, which is like a plasmid, but it capable of containing much larger pieces of DNA, up to 50 kb compared to about 10 kb in a plasmid. Like plasmids, fosmids are circular. However, unlike plasmids, E.Coli can only ever consume (and therefore replicate) one fosmid, which yields a much lower copy number when cloning. We can get larger pieces of DNA, but less of them, with fosmids vs. plasmids.
-
Organisms have many genes, but they are not always "turned on," or expressed. Changes in an organism's environment can affect which genes are turned "on" or "off." For microorganisms the expression of a specific gene may occur within minutes or even seconds following exposure to a given environmental change. For example, a large group of proteins, known as heat shock proteins (HSPs), were found to be over-expressed following exposure of E. coli cells to a temperature increase of only 5 degrees C. The HSP mRNA is expressed, which then codes for the proteins that help the cell deal with the heat exposure (HSPs degrade proteins that are denatured, and protect ones that are needed).
-
The study of the entire DNA sequence (all of the genes present) in an organism or group of organisms.
-
Large contigs are long pieces of DNA (20,000-150,000 bases long - contig originates from contiguous - meaning one piece), while libraries are a form of archiving the large pieces of DNA. When a library is constructed, individual contigs are cloned into individual Escherichia coli (E. coli) cells, which grow up into colonies on agar plates when plated. Each colony contains many cells, though all those cells are "clones," since they are the result of multiple cellular divisions of the same cell. We select these clones (manually, or these days with robots), and place them into multi-well plates that can archive 96-384 different clones at a time.
-
This is an incredibly abundant group of microorganisms inhabiting the worlds' ocean - not often thought of as an extreme environment. They are most commonly found in deep sea waters from ~150m (450 ft), all the way to the sea-floor (1000-4000 m), where they can make up from 10-40% of the cells found (Karner, M.B., E.F. DeLong, and D.M. Karl 2001. Archaeal dominance in the mesopelagic zone of the Pacific Ocean. Nature 409:507-510). This distribution makes it somewhat difficult to study them. Further complicating their study is the fact that we haven't figured out how to culture them yet.
-
Technology developed in the early 1990's which adapts previous DNA hybridization approaches to a microscale format in which the DNA for thousands of genes can be spotted on a substrate (a microscope slide, silica chip or other surface) in a grid (array) in which the address of each gene is known. Nucleic acids (mRNA converted to cDNA, or DNA itself) from samples of interest are then labeled with fluorescent dyes and allowed to find perfect matches with genes on the array, the process of hybridization. Microarrays are most commonly used to study gene expression - to visually detect which genes in an organism or group of organisms are "turned on" (expressed) or "turned off" (latent).
- Open reading frame (ORF)
- Regions in prokaryotic genomes that code for proteins are called open reading frames, or ORFs. An ORF is a DNA fragment whose 5' end starts at a start codon, has a series of consecutive triplets (codons) and ends at a stop codon. All protein-coding genes are contained in ORFs, but not all recognized ORFs contain proteins.
-
Cold-loving organisms that are uniquely adapted to cold water, sediments, soils, snow, and ice, and grow optimally at temperatures less than 10°C.
-
The goal of the shotgun arrays is to identify functional genes of interest directly from gene expression/microarray hybridization experiments (those that have "interesting" differential expression signals, are highly expressed etc.), and thereby circumvent the sequencing step, which is time consuming and costly. The shotgun libraries are prepared, then the inserts from the random libraries are amplified by PCR, purified, and spotted directly on the microarrays. These are called shotgun arrays, since though we know which subclone the spots come from, we do not know anything other than the phylogenetic identity of the genome from which the subclones were generated.
-
DNA is sheared randomly by partial digestion with restriction endonucleases, mechanical shearing by forcing through a small bore needle, or as in our case a process called nebulization. The ends of the randomly sheared DNA are treated to make them "clean" or blunt ended - then these fragments are ligated into plasmids which are transformed into E. coli, then picked into their own 96-well plates and processed to sequence the DNA. In our application we have prepared shotgun libraries of already cloned large-contig DNA (from the APEL), since it's very difficult to sequence directly off of the fosmid DNA as a template (it's too large, and difficult to get in high enough concentrations). The plasmids are in high copy number in each E. coli cell, making it easy to get high yields of plasmid DNA from each clone.
-
The concentration of salts present in ocean water results in a depression of the freezing point. That is, the salts present interrupt the formation of the ice crystal structure of water, and lower (depress) the temperature at which water freezes.
