Geochemistry & Ecology of Red Mat Systems (GERMS)

Undergraduate Summer Research Program

Week Three

Microscopy, Enumeration, and Digital Imaging:

Mat Layer Processing:

Site: Hillside, Red Layer

Image – Visible Light

10X	40X

Image – F Fluorescence

10X	40X

Image – R Fluorescence

10X	40X

Grid-Based Counts:

Replicate 1	Replicate 2
No filaments found	No filaments found

Chemical Analysis of Acid-Digested Samples for Metal Assessment

Specific Post-Acid Digestion Results: Site: Firehole River

Copper	0.01mg/L Clear color
Aluminum	0.00mg/L Pink-orange color
Iron	0.36mg/L Purple color
Zinc	0.09mg/L Orange color
Manganese	Test not performed

Pigment Analysis

Methanol Extraction: Site: Hillside

Methanol Extraction Procedure: Methanol extraction is a process of isolating the pigment in the mat. Using methanol will separate the pigment from the protein and the centrifuge will separate them in the solution so that the pigment in the supernatant can be analyzed.
Methanol Extraction Results: Using a spectrophotometer to read the wavelengths of the red layer mat sample I found four peaks. Two are significant, between 600 and 800nm. The pigments in my sample contain Bacteriochlorophyll a and c at 765nm and 667nm; and have the Phylogenetic Lineage of Purple/Proteobacteria and Green Sulfur Bacteria.

In vivo Bead-Beating: Site: Hillside

In vivo Bead-Beating Procedure: Using a special BB tube and .1mm zirconium beads we placed our sample with necessary reagents in the bead-beating machine to break the membrane of the cells apart and separate the protein and pigment from the membrane.
In vivo Bead-Beating Results: The results of the spectrophotometer analysis are that there is Bacteriochlorophyll c at 796nm and Bacteriochlorophyll a at 879nm in the sample. These results match the results of the methanol pigment analysis.