Introduction to Discover Biology Lab and BIOMES Chambers

For Discover Biology Lab

Welcome to Discover Biology Lab.  Each week, I will have a reading assignment for you.  Because there is no textbook requirement for this course, the reading assignments will typically be in the form of blog posts on my Life and Biology website.  There will be quizzes on the material discussed in the reading assignment, so it is in your best interest to read them!

Biology is a world-wide phenomenon.  We are set to discover many different topics of biology in an introductory manner, so covering the whole world is a bit out of the question.  Instead, we will use the concept of ecosystems as our means of discovering biology. 

Now, ecosystem is a bit of a “fuzzy” word because an ecosystem is defined by the person that wants to study it.  Ecosystems are essentially just systems, so we need to think in terms of systems.  What is a system?  A system is a defined area that is either “closed” or “open”.  In a closed system, nothing gets in or out of the defined space.  By “nothing”, I mean matter and energy are confined within the system.  When we learn about calorimetry later in the semester, we will think through the advantages of using a closed system. 

Ecosystems tend to be open.  That is, energy comes into the ecosystem, moves through the system, and then leaves the system.  Same goes with matter.  The ecosystem we will use in the lab this semester is a “BIOMES chamber”.  This chamber is an enclosure that I artificially designed based on the cost-cutting measure of creating two such enclosures from a single sheet of “blue board” (a plywood-sized sheet of Styrofoam that is two inches thick).  The size of the chamber also works well for the 1-foot square light source in the chamber.  Moreover, these chambers can fit beneath the work benches in the teaching lab.

Let’s get back to the idea of the BIOMES chamber.  First off, BIOMES is an acronym for “Biology of Indoor Organismal and Microbial Ecosystem Sustainability.”  Let’s break that down further.  B is for biology, the study of life.  We can have arguments later to discuss what life is.  Next up in the acronym is I for indoor.  Why indoor?  Well, because we live in Butte and the outdoors is a rather cold place most of the year.  Furthermore, we will house our BIOMES chambers within the teaching lab.  OM means organismal and microbial.  By evaluating plant, animals, and single-celled microbes in our BIOMES chambers, we will span a wide range of life forms all in a little box.  Ecosystems have been discussed above, but don’t worry, we’ll talk more about those soon!  Finally, the S stands for sustainability.  Ecosystems devoid of human interference are often sustainable and we will look at concepts behind those ecosystems to create our human-controlled ecosystems.

What are some of the benefits to using BIOMES chambers this semester?  Previously, experiments in the Discover Biology Lab were haphazard and did not overlap from one week to the next.  There was an effort to match the current topic in class with the experimental topic for that week.  However, we will approach the lab portion of this class as a semester-long project.  We will learn about the scientific method and experimental design.  With these tools, we will then create a series of experiments that will address the overarching goal of finding the most efficient method for growing an indoor crop.  Most projects are constrained by time commitment and costs, so we will factor in these concerns as we proceed. 

See you in class!

Why work with Escherichia coli (E. coli)?

For Discover Biology Lab (Background Information)

It would probably be difficult to find a molecular biology laboratory that didn’t have a freezer full of E. coli.  Species that are commonly used in biology labs are called model organisms.  You will find that there are people that can cite numerous reasons why one particular organism is a “bad” model organism but, in practice, there is no perfect model organism.  This essay will describe some of the reasons why scientists use E. coli.

Ease and cost are two factors that should be a considered when working in a lab.  E. coli excels at both of these.  The first two talking points (figure 1) are the facts that, like all bacteria, they reproduce by binary fission and, unlike all bacteria, they can do so very rapidly.  Under optimal conditions, E. coli can replicate about once every 20 minutes.  In contrast, Mycobacterium tuberculosis has a much slower optimal growth rate (1).

The final talking point in this section is that the medium (typically Luria-Bertani broth or “LB” broth) is both easy to make and inexpensive.  The ingredients are 10 grams tryptone, 10 grams sodium chloride (table salt), and 5 grams yeast extract per liter of water (2).  Add some E. coli to LB broth, heat it up to 37 degrees Celsius (aka body temperature or 98.6 degrees Fahrenheit) and shake it at about 200-220 rpm to aerate the sample and the bacteria will multiply for you.

Figure 1: Benefits of working with E. coli (part 1)

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Bacteria replicate by  binary fission (figure 2).  This means that one “parent” cell will divide into identical “daughter” cells.  As the bacteria continue to replicate, their population size will repeatedly double.  A generic growth curve is shown in in figure 3. At first the doubling rate is slow and this represents the “lag” period of growth.  Next, the replication is optimal and this growth phase is referred to logarithmic (or “log”) growth.  As nutrients become limiting, the growth rate slows to a standstill (stationary phase).  After this, the bacteria will begin dying faster than replicating and the population will crash.

Figure 2: Replication by binary fission (one becomes two)

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Figure 3: Typical growth curve

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When certain species of bacteria are placed in a stressful environment, such as higher than normal temperatures, they can increase their activity of taking up DNA from the environment through a process called transformation.  E. coli can be grown in such a manner as to increase the competence of taking up DNA in response to stress.  In fact, we follow a protocol for making competent bacteria to create an inexpensive source of these in my lab.  The extracellular DNA that is picked up from the environment can be in the form of plasmid DNA (figure 4).

Figure 4: Chromosomal (genomic) and extrachromasomal (plasmid) DNA in E. coli

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Like genomic DNA chromosomes, plasmid DNA is circular.  Each plasmid has an origin of replication (aka “ori”) that will recruit the same DNA replication machinery used in replication of genomic DNA.  Depending on the sequence in the origin of replication, the plasmids will be found at varying concentrations in each bacteria.

Many cellular and molecular biology research tools are built from plasmids that were created and modified for a particular purpose.  Common and useful genetic elements in a variety of plasmids will be covered in a future post.

Figure 5: Benefits of working with E. coli (part 2)

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In conclusion, it’s the ability of E. coli to make copies of plasmids is what makes them such a welcomed addition to most biology laboratories.  The last aspect to discuss in this post is the ability to easily and cheaply store these plasmids and plasmid makers (figure 6).  Monocultures of unique strains of bacteria that copy a specific plasmid can be stored in a 50-50 glycerol/growth medium mix at very cold temperatures (typically -80C).  Amazingly, these “frozen” cultures can be stored for years to decades in this state.

Alternatively, cultures of bacteria can be busted open and, through a plasmid preparation procedure, the plasmid DNA can be purified.  The plasmid DNA is usually stored in water (or Tris-EDTA buffer) at -20C for months to years.

Finally, if you’d like to ship some plasmid DNA through the mail to a collaborator, just drop some plasmid solution onto heavy weight paper and let it dry.  If you circle where the plasmid dried, your collaborator can cut out that area of the paper and suspend the DNA in some water.  A quick bacterial transformation will produce plasmid makers at the destination site.

Figure 6: Benefits of working with E. coli (part 3)

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References

  1. http://mmbr.asm.org/content/72/1/126.full
  2. http://cshprotocols.cshlp.org/content/2006/1/pdb.rec8141.full?text_only=true