Megan's Notes

Notes on Thermus Aquaticus

1. Hot Springs (Yellowstone)
   1. example of biodiversity in an extreme enviorment
   2. number of species of bacterium quite small, yet distinct
      1. Thermus Aquaticus
         1. isolates Taq polymerases
            1. heat resistant enzyme crucial for DNA amplification used in research and medical diagnoes

Interesting Facts

1. In the 1980s an enzyme from the bacteria-like Thermus aquaticus became the key to a technique of sequencing DNA that made possible the work of the Human Genome Project, which produced a genetic map of the body, and is today a cornerstone of the $300-billion-a-year biotech industry.

2. Awesome Video~http://school.nettrekker.com/tts/external.epl/http://highered.mcgraw-hill.com/olc/dl/120078/micro15.swf

 

Classification

1. Extremophiles
   1. named for there ability to survive in enviorments in Yellowstone National Park that include gysesrs and mud pots

History

1. Discovered in 1969
   1. Thomas D. Brock
      1. derialed the theory that life couldn't exist in temperatures above 163 degrees F

Beneficial Properties

The following uses the Polymerare Chain Reaction to benefit society:

1.     Criminal Investigation
   1. helps investigators work quickly and easily with DNA evidence
      1. Example
      2. September 11th, 2001 identified those who died in the Pentagon and World Trade Centers

1.       Doctors
   1. use the process of DNA sequence to diagnose and treat genetic disease

1.         Park Service 
   1. PCR helps Parks Servise identify and manage park wildlife

1. Reseachers
   1. use the process to learn more about other extremeophiles
   2. use to make billions of copies of DNA sequences in a few hours

Polymerase Chain Reaction

1.  a method of copying genes and a crucial step in DNA analysis
   1. scientists were able to extract an enzyme from Thermus Aquaticus that remained stable at near boiling temperatures

 

Name Thermus Aquaticus

1.   Derived from latin

a.   Aqua- water

2.   Derived from Greek

a.   Thermos- hot

3.   T.  Aquaticus is an aquatic thermophile

Structure and Form

1.   Rods and Filaments

2.   Gram-negative

3.   Flagella and endospores absent

4.   Nonsporulating

5.   Nonmotile

6.   Rods

a.   Frequently form long filaments

                                         i.    At superoptimal temperatures

                                        ii.    Stationary phase

7.   All isolates form a large sphere

a.   Larger than a spheroplast

b.   Resistant to osmotic lysis

Colony Characteristics

1.   Compact, slowly spreading colonies

2.   Yellow to bright orange on yeast extract plus mineral salts-agar

Liquid Culture

1.   Yellow surface pellicle often formed in unshaken liquid cultures

Nutrition

2.   No growth factor requirements

3.   Thrives on

a.   Nitrogen sources

b.   Amino acids

c.   NH₄ +

d.   Carbon sources

e.   Sugars

f.    Organic acids

4.   Grows best in complex media such as 0.1-0.3% tryptone yeast extract

5.   Did not require vitamins or amino acid for growth

a.   Grows faster in an enriched environment

6.   Several sugars and organic acids are the carbon source

7.   Either NH₄ of glutamate could be nitrogen source

Oxygen

1.   Obligately aerobic

pH levels

2.   Optimum 7.5 to 7.8

3.   No growth below pH 6 or above pH 9.5

Morphology

1.   Isolates are filamentous

a.   During continued transfer the filaments become shorter

2.   Influenced by temperature of growth

3.   Influences by growth stage of the culture

Misc.

1.   Rapidly destroyed by sodium lauryl sulfate

2.   Resistant to osmotic lysis

a.   Due to unusually strong cell membrane of t.a.

Habitat

2.   Strains of T. Aquaticus isolated in

a.   thermal springs in Yellowstone National Park

b.   thermal springs in California

c.   Man Made Thermal Habitats

                                         i.    Hot tap water far from thermal springs

3. http://streaming.discoveryeducation.com/search/assetDetail.cfm?guidAssetID=0660A56E-52BC-4EFF-9331-031E3A79B386 (video clip about Yellowstone)

Growth

1.   The growth of all isolates is inhibited by a fairly low concentration of:

a.   Cyloserine

b.   Streptomycin

c.   Penicillin

d.   Novobiocin

e.   Tetracycline

f.    Chloramphenicol

Polymerase Chain Reaction

1. The numbers of species found in these places may be smaller than almost anywhere else, yet the species are quite distinctive. One such species is the bacterium Thermus aquaticus, found in the hot springs of Yellowstone. From this organism was isolated Taq polymerase, a heat-resistant enzyme crucial for a DNA-amplification technique widely used in research and medical diagnostics (see polymerase chain reaction).

1. developed in 1983 by Kary B. Mullis
2. prior to discovery
   1. methods to amplify or generate copies of DNA was time-consuming and labor intensive.
   2. now a machine that carries out PCR uses an enzyme from T.a. to replicate billions of DNA in a few hours

• three step process in repeated cycles

1. denaturation, seperation of the DNA molecule (accomplished by heating at 203 degrees F
2. temperature is reduced to 131 degrees F so that primers, short stretches of nucleotides that correspond to the template sequence, can anneal to the template
3. the temperature is raised to 162 degrees F and the DNA polymerase begins adding nucleotides onto the ends of annealed primers

1. process repeated 25-30 times to make a lot of DNA

• problems without T.A.

1. the DNA polymerase had to be replenished after every cycle because it is not stable at the high temperatures needed for denaturation
2. problem solved in 1987 with the discovery of a heat-stable polymerase called Taq, an enzyme isolated from the thermophilic bacterium Thermus aquaticus,
3. Taq polymerase led to the invention of the PCR machine

Quotes

1. "The PCR process is lighting [scientists] path as they move through these projects," says Susan Kelly, outreach and education coordinator from the Thermal Biology Institute at Montana State University. "