RESEARCH 2002
RESEARCH 2001
RESEARCH 2000

Jenny Strickland
Purdue University
Mentor: Dr. James Maki

Yellowstone National Park is one of the most exceptional areas in the world. A magma chamber about 5 miles under the park creates many distinctive features. Because of the high amount of hydrothermal activity in the area many unique ecosystems are found. One of these ecosystems is seen in Yellowstone Lake. Yellowstone Lake is the largest high altitude lake in North America at relatively 2000 meters above sea level. The northern and major part of the lake, including Mary bay, is part of the Yellowstone caldera. This caldera was formed when a magma chamber collapsed during a volcanic explosion over 600,000 years ago. The location of the lake in this caldera causes a unique topology on its floor. Many hydrothermal vents and springs are found in the lake and are of high interest to scientists. 

The Maki lab studies the microbial make-up of the water that is emitted from the vents in Yellowstone Lake. From their research they have discovered that methanotrophic bacteria are present in water samples collected. Methanotrophic bacteria use methane as a carbon and energy source. These methane-utilizing bacteria have an enzyme called methane monooxygenase that oxidizes CH4 to CO2. There are two forms of this enzyme; sMMO and pMMO. The pMMO form is of interest in our research. This form is found in all methanotrophs and is a membrane bound enzyme. All methanotrophs have a gene construct for pMMO named pmoCAB. PmoA is much conserved, approximately 330bp, and has two copies in the genome. This gene is targeted for detection of methanotrophs in water samples from Yellowstone Lake.

Previously amplified samples 99-13-1 and 02-11-stb were cloned into E.coli using an Invitrogen cloning kit and grown on LB plates containing carbanicillin and x-gal. The white colonies were then used for direct colony PCR. Previously constructed pmoA specific primers were used to amplify the samples with a PCR protocol provided by Jim Bruckner. The samples were then run on a 1% agarose gel to check positive pmoA detection. The positive samples were then digested with a double digest of MspI and HaeIII. Further, a single digest was performed with more of the sample using HhaI. The digests were then run next two each other on a 2% agarose gel. Fragment patterns were reviewed and vent profiles were made. In the future, distinct vent samples will be sent in for sequencing, and identification using a genetic database will be preformed.