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Studied by 25 peopleHost Basics-Arthrobacter sp.
An Introduction to the Host Bacteria, Arthrobacter sp.
As a network of researchers, SEA-PHAGES explores the genetic diversity and relationships of populations of bacteriophages that infect Actinobacteria, incluidng the bacterium Arthrobacter sp. There are many different strains of Arthrobacter sp. (isolated from different environmental samples). The Arthrobacter sp. strain widely used in SEA-PHAGES is called Arthrobacter sp. KY3901. This strain is from the American Type Culture Collection (catalog # 21022). If you intend to isolate phages using this bacterium, then this will be your ‘host’ for phage isolation.
Arthrobacter sp. is one of over 70 species that make up the genus Arthrobacter, classified within the phylum Actinobacteria. Arthrobacter sp. is a gram-positive soil organism, whose relatives are well equipped to breakdown various hydrocarbons. Arthrobacter arilaitensis, for instance, grows on the surface of cheese and catabolizes the fatty acids, amino acids, and lactic acid present, contributing to the color and flavor of the cheese. Other Arthrobacter species are capable of breaking down compounds such as hexavalent chromium, which is a carcinogen, suggesting a potential use in bioremediation (i. e. the breakdown of pollutants). The specific Arthrobacter strain you will use, alongside its relatives, has been found to produce penicillin derivatives and aspartic decarboxylase.
Arthrobacter sp. will grow and divide on many nutrient-rich media. In your research, you will grow Arthrobacter sp on PYCa media and at 22 - 28˚C. Under these conditions, Arthrobacter sp grows rapidly, dividing approximately once every 2 - 3 hours. This means that it typically takes about 3 days for an individual cell to form a colony on an agar plate. However, when preparing a bacterial
lawn, you can start by adding many bacterial cells (e. g. 107 or more) so that a lush lawn grows within 1 – 2 days. For reproducibility of experiments, it is important that cultures used for a given set of experiments be prepared at the same temperature.
Arthrobacter sp colonies are tan to yellow in color, smooth and glossy (Figure 4.0-4). When incubated for prolonged periods, the color of the colonies intensifies. A liquid culture of Arthrobacter sp inoculated from a single colony will take 1 – 2 days to form a saturated liquid
culture, exhibiting a tan color. The strain you will work with, Arthrobacter sp. KY3901, is not inhibited by the antifungal cyclohexamide or the antibiotic spectinomycin. Therefore, this can be added to the growth media to prevent other microorganisms from growing in your cultures.
On average, 40 % of attempts to isolate phage from soil by enrichment with Arthrobacter sp. yield phage
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Figure 4.0-4. Arthrobacter sp. growing on an agar plate. |
If you are using this bacterium as your host for phage isolation, refer back to the list below for the specific growth and culture requirements for your experiments.
Growth media: PYCa
Growth temperature: 22 ˚C - 28˚C (this host does not grow at 37˚C)
Antimicrobials: cycloheximide (10 μg/ml), spectinomycin (optional)
Phage Buffer: 10 mM Tris (pH 7.5), 10 mM MgSO4, 68 mM NaCl, 1 mM CaCl2, (10% glycerol, optional)
Restriction Enzymes: BamHI, ClaI, EcoRI, HaeIII, HindIII, MseI
Note: Isoschizomers, which are different restriction enzymes that recognize the same DNA sequence, may be used in place of any of the enzymes listed above.